HCV NS-3 serine protease inhibitors

ABSTRACT

Methods drawn to peptidomimetic compounds which inhibit the NS3 protease of the hepatitis C virus (HCV), are described. The compounds have the formula (VI) where the variable definitions are as provided in the specification. The compounds comprise a carbocyclic P2 unit in conjunction with a novel linkage to those portions of the inhibitor more distal to the nominal cleavage site of the native substrate, which linkage reverses the orientation of peptidic bonds on the distal side relative to those proximal to the cleavage site.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.12/642,984, filed Dec. 21, 2009, which is a divisional of U.S.application Ser. No. 10/572,349, filed Jan. 2, 2007, which is theNational Stage filing of International Application No.PCT/SE2005/000097, filed Jan. 28, 2005, which claims the benefit ofSE0400199-6, filed Jan. 30, 2004, SE 0401288-6, filed May 19, 2004, andSE 0402562-3, filed Oct. 22, 2004, the entireties of which areincorporated by reference herein.

TECHNICAL FIELD

This invention relates to novel inhibitors of the NS3 serine protease ofthe flavivirus HCV and to methods for their use in the treatment orprophylaxis of HCV.

BACKGROUND ART

The NS3 serine protease of HCV is a multifunctional protein whichcontains a seine protease domain and a RNA helicase domain. The proteasecofactor NS4A, which is a relatively small protein, is absolutelyrequired for enhanced serine protease activity. The NS3 serine proteaseis essential in the viral lifecycle. From analysis of the substratebinding site as revealed by X-ray crystal structure, it has been shownthat the binding site of the NS3 protease is remarkably shallow andsolvent exposed making small molecule inhibitor design a challenge.

It is believed that two HCV protease inhibitors have entered clinicaltrials, namely Boehringer Ingelhelm's BILN-2061 disclosed in WO 0059929and Vertex′ VX-950 disclosed in WO 0387092. A number of similarpeptidomimetic HCV protease inhibitors have also been proposed in theacademic and patent literature. Common for the vast majority of suchprior art peptidomimetics is the presence of an L-proline derivative atthe P2 position of the inhibitor and interacting with the S2 subsite ofthe HCV protease enzyme. In the case of BILN-2061, the L-proline is4-substituted with a quinoline ether, whereas VX-950 has a carboyclicring fused to the L-proline ring. Most peptidomimetics additionallycomprise additional L-amino acid derivatives peptide bonded at the P3position, with many proposed inhibitors also including additionalL-amino acid derivatives extending into P4, P5 and P6.

It has already become apparent that the sustained administration ofBILN-2061 or VX-950 selects HCV mutants which are resistant to therespective drug, so called drug escape mutants. These drug escapemutants have characteristic mutations in the HCV protease genome,notably D168V, D168Y and/or A165S. Treatment paradigms for HCV will thushave to resemble HIV treatment, where drug escape mutations also arisereadily. Accordingly, additional drugs with different resistancepatterns will consistently be required to provide falling patients withtreatment options, and combination therapy with multiple drugs is likelyto be the norm in the future, even for first line treatment.

Experience with HIV drugs, and HIV protease inhibitors in particular,has further emphasized that sub-optimal pharmacokinetics and complexdosage regimes quickly result in inadvertent compliance failures. Thisin turn means that the 24 hour trough concentration (minimum plasmaconcentration) for the respective drugs in an HIV regime frequentlyfalls below the IC₉₀ or ED₉₀ threshold for large parts of the day. It isconsidered that a 24 hour trough level of at least the IC₅₀, and morerealistically, the IC₉₀ or ED₉₀ is essential to slow down thedevelopment of drug escape mutants and achieving the necessarypharmacokinetics and drug metabolism to allow such trough levelsprovides a stringent challenge to drug design. The stronglypeptidomimetic nature of prior art HCV protease inhibitors, withmultiple peptide bonds in native configurations poses pharmacokinetichurdles to effective dosage regimes.

BRIEF DESCRIPTION OF THE INVENTION

In accordance with a first aspect of the invention, there are providedcompounds of the formula VI:

whereinA ss C(═O)OR¹, C(═O)NHSO₂R², C(═O)NHR³, or CR⁴R^(4′) wherein;R¹ is hydrogen, C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl,C₀-C₃alkylheterocyclyl;R² is C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl;R³ is C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl,—OC₁-C₆alkyl, —OC₀-C₃alkylcycarbocyclyl, —OC₀-C₃alkylheterocyclyl;R⁴ is halo, amino, or OH; or R⁴ and R^(4′) are ═O;R^(4′) is C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl;whereinR², R³, and R^(4′) are each optionally substituted from 1 to 3substituents independently selected from the group consisting of timeswith halo, oxo, nitrile, azido, nitro, C₁-C₆alkyl,C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl, NH₂C(═O)—, Y—NRaRb,Y—O—R_(b), Y—C(═O)Rb, Y—(C═O)NRaRb, Y—NRaC(═O)Rb, Y—NHSO_(p)Rb,Y—S(═O)_(p)Rb, Y—S(═O)_(p)NRaRb, Y—C(═O)Orb and Y—NRaC(═O)ORb;Y is independently a bond or C₁-C₃alkylene;Ra is independently H or C₁-C₃alkyl;Rb is independently H, C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl orC₀-C₃alkylheterocyclyl;p is independently 1 or 2;M is CR⁷R^(7′) or NRu;Ru is H or C₁-C₃alkyl;R⁷ is C₁-C₆alkyl, C₀-C₃alkylC₃-C₇cycloalkyl, or C₂-C₆alkenyl, any ofwhich is optionally substituted with 1-3 halo atoms, or an amino, —SH orC₀-C₃alkylcycloalkyl group, or R⁷ is J;R^(7′) is H or taken together with R⁷ forms a C₃-C₆cycloalkyl ringoptionally substituted with R^(7′a) wherein;R^(7′a) is C₁-C₆alkyl, C₃-C₅cycloalkyl, C₂-C₆alkenyl any of which may beoptionally substituted with halo; or R^(7′a) is J;q′ is 0 or 1 and k is 0 to 3;Rz is H, or together with the asterisked carbon forms an olefinic bond;Rq is H or C₁-C₆alkyl;W is —CH₂, —O—, —OC(═O)H—, —OC(═O)—, —S—, —NH—, —NRa, —NHSO₂—,—NHC(═O)NH— or —NHC(═O)—, —NHC(═S)NH— or a bond;R⁸ is a ring system containing 1 or 2 saturated, partially saturated orunsaturated rings each of which has 4-7 ring atoms and each of which has0 to 4 hetero atoms selected from S, O and N, the ring system beingoptionally spaced from W by a C₁-C₃alkyl group; or R⁸ is C₁-C₆alkyl; anyof which R⁸ groups can be optionally mono, di, or hi substituted withR⁹, whereinR⁹ is independently selected from the group consisting of halo, oxo,nitrile, azido, nitro, C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl,C₀-C₃alkylheterocyclyl, NH₂CO—, Y—NRaRb, Y—O—Rb, Y—C(═O)Rb,Y—(C═O)NRaRb, Y—NRaC(═O)Rb, Y—NHSO_(p)Rb, YS(═O)_(p)Rb,Y—S(═O)_(p)NRaRb, Y—C(O)Orb and Y—NRaC(═O)ORb; wherein said carbocyclylor heterocycyl moiety is optionally substituted with R¹⁰; whereinR¹⁰ is C₁-C₆alkyl, C₃-C₇cycloalkyl, C₁-C₆alkoxy, amino, sulfonyl, (C₁-C₃alkyl)sulfonyl, NO₂, OH, SH, halo, haloalkyl, carboxyl, amido,Rx is H or C₁-C₅ alkyl; or Rx is J;T is —CHR¹¹— or —NRd-, where Rd is H, C₁-C₃alkyl; or Rd is J;R¹¹ is H, C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl, anyof which can be substituted with halo, oxo, nitrile, azido, nitro,C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl, NH₂CO—,Y—NRaRb, Y—O—Rb, Y—C(═O)Rb, Y—(C═O)NRaRb, Y—NRaC(O)Rb, Y—NHSO_(p)Rb,Y—S(═O)_(p)Rb, Y—S(═O)_(p)NRaRb, Y—C(═O)ORb, Y—NRaC(═O)ORb; or R¹¹ is J;J, if present, is a single 3 to 10-membered saturated or partiallyunsaturated alkylene chain extending from the R⁷/R^(7′) cycloalkyl, orfrom the carbon atom to which R⁷, is attached to one of Rd, R, Rx, Ry orR¹¹ to form a macrocycle, which chain is optionally interrupted by oneto three heteroatoms independently selected from: —O—, —S— or —NR¹²—,and wherein 0 to 3 carbon atoms in the chain are optionally substitutedwith R¹⁴; wherein;R¹² is H, C₁-C₆ alkyl, C₃-C₆cycloalkyl, or C(═O)R¹³;R¹³ is C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl;R¹⁴ is independently selected from the group consisting of H,C₁-C₆alkyl, C₁-C₆haloalkyl, C₁-C₆alkoxy, hydroxy, halo, amino, oxo, thioand C₁-C₆thioalkyl;m is 0 or 1; n is 0 or 1;U is ═O or is absent;R¹⁵ is H, C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl, anyof which can be substituted with halo, oxo, nitrile, azido, nitro, C₁-C₆alkyl, C₀-C₃alkylheterocyclyl, C₀-C₃alkylcarbocyclyl, NH₂CO—, Y—NRaRb,Y—O—Rb, Y—C(═O)Rb, Y—(C═O)NRaRb, Y—NRaC(═O)Rb, Y—NHS(═O)_(p)Rb,Y—S(═O)_(p)Rb, Y—S(═O)_(p)NRaRb, Y—C(═O)ORb, Y—NRaC(═O)ORb;G is —O—, —NRy-, —NRjNRj-;Ry is H, C₁-C₃ alkyl; or Ry is J;one Rj is H and the other Rj is H or J;R¹⁶ is H; or R¹⁶ is C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl,C₀-C₃alkylheterocyclyl, any of which can be substituted with halo, oxo,nitrile, azido, nitro, C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl,C₀-C₃alkylheterocyclyl, NH₂CO—, Y—NRaRb, Y—O—Rb, Y—C(═O)Rb,Y—(C═O)NRaRb, Y—NRaC(═O)Rb, Y—NHSO_(p)Rb, Y—S(═O)_(p)Rb,Y—S(O)_(p)NRaRb, Y—C(═O)ORb, Y—NRaC(═O)ORb;or a pharmaceutically acceptable salt or prodrug thereof.

Without in any way wishing to be bound by theory, or the ascription oftentative binding modes for specific variables, the notional conceptsP1, P2, P3, and P4 as used herein are provided for convenience only andhave substantially their conventional meanings, as illustrated bySchechter & Berger, (1976) Biochem Biophys Res Comm 27 157-162, anddenote those portions of the inhibitor believed to fill the S1, S2, S3and S4 subsites respectively of the enzyme, where S1 is adjacent thecleavage site and S4 remote from the cleavage site. Regardless ofbinding mode, the components defined by Formula VI etc are intended tobe within the scope of the invention. For example it s expected thatcapping group R¹⁶-G may interact with the S3 and S4 subsites especiallywhen m and/or n is 0.

The various embodiments of the present invention can be notionallyrepresented as R¹⁶-G-P4-P3-P2-P1, wherein P3 and/or P4 may be absent P1,P3 and P4 each represents a building block constituted of a derivativeof a natural or unnatural amino acid, P2 is a substituted carbocydclicresidue and G-R¹⁶ is a capping group. The building blocks are typicallylinked together by amide bonds which are reversed relative to each otheron each side of the P2 building block in the compounds of the invention.

Additional aspects of the invention include a pharmaceutical compositioncomprising a compound of the invention as defined above and apharmaceutically acceptable carrier or diluent therefor.

The compounds and compositions of the invention have utility in methodsof medical treatment or prophylaxis of HCV infections in humans.Accordingly, a further aspect of the invention is the use of a compoundas defined above in therapy such as the manufacture of a medicament forthe prophylaxis or treatment of flavivirus infections in humans oranimals. Exemplary flavivirus include BVDV, dengue and especially HCV.

In the compounds of the invention the amide bond linking the P2 and P3together is reversed relative to the amide bond linking the P1 and P2,i.e. the amino acid derivatives, P1 and P3, on each side of the P2scaffold are both coupled through their amino functions to the acidgroups on each side of the P2 scaffold. This means that the side chainsof the P3 and P4 (including the R¹⁶ cap to the extent this interactswith S3 or S4) point in the opposite direction compared to in a nativepeptide substrate. Another consequence of the reversed P3 and P4 aminoacids is that the side chains of these amino acids are displaced oneatom outwardly relative to a native peptide substrate.

Change of direction of the P3 and P4 side chains in this fashion wouldbe expected to favour non-natural D stereochemistries for the pocketfiling groups (eg side chains) of P3 and/or P4 and/or R¹⁶. Indeed, suchcompounds are typically highly active and within the scope of theinvention. However, it has been surprisingly found that even compoundsof the invention bearing L-amino acid side chains at P3 and/or P4exhibit good activity, notwithstanding that the respective entity mustapproach the S3 or S4 pocket from a different angle relative to a nativepeptide substrate. Accordingly L-stereochemistry at R¹¹ and/or R¹⁵and/or the corresponding configuration at R¹⁶ to mimic L stereochemistryrepresents a favoured aspect of the invention.

The different angle of approach to the S3 and/or S4 pockets also hasimplications for the ability of the compounds of the invention to avoidresistance patterns exhibited by prior art HCV protease inhibitors whichhitherto have all had a conventional peptide backbone of natural ornon-natural L-amino acid residues. As with the reverse transcriptase ofHIV which is notorious for quickly generating drug escape mutants underthe selective pressure of antiviral therapy, the RNA dependent RNApolymerase NS5A of HCV has a very poor proof reading capacity. This inturn means that the HCV polymerase is highly error prone and it islikely that characteristic resistance patterns will arise when HCVantivirals are administered over long periods. Even before launch, it isapparent that BILN 2061 with a substantially peptidic backbone (albeitmacrocyclised) and Vertex′ NS3 protease inhibitor VX-950 with a linearpeptide backbone at P3 and P4 quickly give rise to characteristicresistance mutations at positions 155, 156 or 168 of the NS3 protease(Lin et al J Biol Chem 2004 279(17):17808-17).

A preferred group of compounds of the invention comprises those whereinP1 represents a hydrazine derivative, that is M is NRu where Ru istypically H or C₁-C₃alkyl. Compounds wherein M is CR⁷R^(7′) constitute afurther preferred aspect of the Invention.

Preferred embodiments wherein M is CR⁷R^(7′) in formulae VI includeformulae VIA below.

Preferred values for q′ and k in formula VI include 1:1, 1:2, 1:3, 22,2:3, more preferably 0:2 and 0:0; and most preferably 0:1, in which casepreferred compounds have one of the partial structures:

especially where Rz is H or Rq is H or methyl.

Compounds of the invention may comprise both a P3 and a P4 function, vizm and n are each 1. Favoured embodiments within Formula VI comprisingboth a P3 and P4 function include formula VIda-VIdb below:

Alternative embodiments include the structures corresponding to VIda,and VIdb wherein M is NRu.

Alternative configurations of the compounds of the invention comprise aP3, but no P4 function, viz m is 1 and n is zero. Favoured embodimentswithin Formula VI comprising a P3, but no P4 include formula VIea-VIebbelow:

Alternative embodiments include the structures corresponding to VIea andVIeb wherein M is NRu.

Still further alternative configurations of the compounds of theinvention include those where m and n are zero and thus groups R¹⁶-Gabut P2, but as mentioned above, the capping group R16-G may interactfavourably with S3 and/or S4.

Favoured embodiments within Formula VI wherein m and n are zero includethose of formula VIfa below:

Alternative embodiments include the structures corresponding to VIfa,wherein M is NRu.

The compounds of the invention may comprise linear molecules, asdepicted above. Alternatively, in embodiments wherein R⁷ and R^(7′)together define a spiro cycloalkyl group, such as spiro-cyclopropyl, thecompounds of the invention may be configured as macrocycles, wherein alinking group J extends between one of Rj, Rx, Ry, Rd or R¹¹ of FormulaVI. Alternatively the macrocycle J may extend from the carbon adjacentR⁷ to one of RJ, Rx, Ry, Rd or Ru.

Favoured embodiments of such macrocyclic structures within formula VIwherein m is 0 and n is 1 include those of the formula VIga-VIgc below:

The corresponding structures wherein the J chain bonds to the carbonadjacent R⁷ are also favoured.

Favoured embodiments of macrocyclic structures within formula VIcomprising both a P3 and P4 functions, i.e. wherein both m and n are 1,include those of the formula VIha-VIhc below:

The corresponding structures wherein the J chain bonds to the carbonadjacent R⁷ are also favoured.

Favoured macrocyclic structures within Formula VI, wherein both of theP3 and P4 functions are absent, i.e. wherein m and n are each 0, includethose of the formulae VIhe-VIhf below.

The corresponding structures wherein the J chain bonds to the carbonadjacent R⁷ are also favoured.

In general, in the optionally macrocyclic structures such as thoseillustrated above, linker J is a 3 to 10 chain atom, preferably 4 to 7chain atom, such as 6 or 6 chain atom, saturated or partiallyunsaturated alkylene chain, that is an alkylene chain bearing 1 to 3unsaturated bonds between adjacent carbons, typically one unsaturation.The length of the chain will, of course, depend on whether J extendsfrom Rd, Rx, Rx, Ry, R¹¹ or the carbon adjacent R⁷. Suitable chains aredescribed in detail in WO 00/59929. Typically J will be dimensioned toprovide a macrocycle of 13 to 16 ring atoms (including those atoms inthe P1, P2 and if present P3 groups contributing to the ring).Conveniently J is dimensioned to provide a macrocycle of 14 or 15 ringatoms.

Conveniently, the J chain contains one or two heteroatoms selected from:O, S, NH, NC₁-C₆ alkyl or N—C(═O)C₁-C₆alkyl. More preferably, the Jchain optionally contains one heteroatom selected from: NH, orN—C(═O)C₁-C₆alkyl, most preferably N(Ac). Most preferably, the chaincontaining a nitrogen atom is saturated. In an alternative embodiment, Jcontains one heteroatom selected from O or S. The chain may besubstituted with R¹⁴, such as H or methyl.

Typically the J linker structure is saturated. Alternatively, J contains1 to 3, preferably one double bond, typically spaced one carbon from thecycloakyl R⁷ function, if present. The double bond may be cis or trans.

Representative examples of J thus include pentylene, hexylene,heptylene, any of which are substituted with C₁-C₆alkyl, C₁-C₆haloalkyl,C₁-C₆alkoxy, hydroxyl, halo, amino, oxo, thio or C₁-C₆thioalkyl;penten-3-yl, hexen-4-yl, hepten-5-yl, where 3, 4 or 5 refers to a doublebond between carbon atoms 3 and 4, 4 and 5 etc.

Convenient R⁷ and R^(7′) groups include those wherein R^(7′) is H and R⁷is n-ethyl, n-propyl, cyclopropyl, cyclopropylmethyl, cyclobutyl,cyclobutylmethyl, 2,2-difluoroethyl, or mercaptomethyl. Preferredembodiments include those wherein R⁷ is n-propyl or 2,2-difluoroethyl.

Alternative favoured configurations for R⁷ and R include those whereinR^(7′) is H and R⁷ is C₃-C₇ cycloakyl or C₁-C₃alkylC₃-C₇cycloalkyl.

Still further favoured configurations for R⁷ and R^(7′) include thosewherein R^(7′) is H and R⁷ is J.

Alternatively, R⁷ and R^(7′) together define a spiro-cycloalkylfunction, such as a spiro-cyclobutyl ring, and more preferably aspiro-cyclopropyl ring. “Spiro” in this context simply means that thecycloakyl ring shares a single carbon atom with the peptidic backbone ofthe compound. The ring is substituted or unsubstituted. Preferredsubstituents include mono or di-substitutions with R^(7′a) whereinR^(7′a) is C₁-C₆ alkyl, C₃-C₆cycloalkyl, or C₂-C₆ alkenyl, any of whichis optionally substituted with halo.

Alternatively the substituent may be a J linker as described above.Currently preferred stereochemistries for a spiro-cyclopropyl ring aredefined below.

Particularly preferred substituents include R^(7′a) as ethyl, vinyl,cyclopropyl (ie a spiro-cyclopropyl substituent to the spiro cycloalkylring of R⁷/R^(7′)), 1- or 2-bromoethyl, 1- or 2-fluoroethyl,2-bromovinyl or 2-fluorethyl.

In one embodiment of the invention A is —CR⁴R^(4′) as illustrated indetail in PCT/EP03/10595, the contents of which are incorporated byreference.

Convenient R^(4′) groups thus include C₁-C₆alkyl, such as methyl, ethyl,propyl, ethenyl and —CHCHCH₃. Alternative preferred R groups includearyl or heteroaryl such as optionally substituted phenyl, pyridyl,thiazolyl or benzimidazolyl or C₁-C₃alkylaryl or C₁-C₃alkylheteroaryl,where the alkyl moiety is methyl, ethyl, propyl, ethenyl and —CHCHCH₃.Preferred aryl moieties include optionally substituted phenyl,benzothiazole and benzimidazole.

Favoured R⁴ groups include —NH₂, fluoro or chloro. Alternative preferredR⁴ groups include —OH and especially ═O.

An alternative embodiment for A is C(═O)NHR³, where R³ is optionallysubstituted C₀-C₃alkylaryl, C₀-C₃alkylheteroaryl, OC₀-C₃alkylaryl orOC₀-C₃alklylheteroaryl. Appropriate substituents appear in thedefinitions section below.

An alternative favoured configuration for A s C(═O)OR¹, especially whereR¹ is C₁-C₆alkyl, such as methyl, ethyl, or tert-butyl and mostpreferably hydrogen.

A particularly preferred configuration for A is C(═O)NHSO₂R², especiallywhere R² is optionally substituted C₁-C₆alkyl, preferably methyl, oroptionally substituted C₃-C₇cycloakyl, preferably cyclopropyl, oroptionally substituted C₀-C₆alkylaryl, preferably optionally substitutedphenyl. Appropriate substituents appear in the definitions sectionbelow.

Substituent —W—R⁸ on the cyclic P2 group can employ any of the prolinesubstituents which are extensively described in WO 00/59929, WO00/09543, WO 00/09558, WO 99/07734, WO 99/07733, WO 02R/0926,WO03/35060, WO 03/53349, WO03/064416, W=03/86103, WO03/064455,WO03/064456, WO03/62265, WO03/062228, WO03/87092, WO 03/99274,WO03/99316, WO03/99274, WO04/03670, WO04/032827, WO04/037855,WO04/43339, WO04/92161, WO04/72243, 5WO04/93798. WO04/93915, WO04/94452,WO04/101505, WO04/101602, WO04/103996, WO04/13365 and the like.

Favoured W functions include W as —OC(═O)NH—, —OC(═O)—, —NH—, —NR^(8′)—,—NHS(O)₂— or —NHC(═O)—, especially —OC(═O)NH— or —NH—. Favoured R⁸groups for such W functions include optionally substitutedC₀-C₃alkylcarbocyclyl or C₀-C₃alkylheterocyclyl, including thosedescribed in WO0009543, WO0009558 and WO 00/174768. For example estersubstituents, —W—R⁸, on the cyclic P2 group, include those disclosed inWO 01/74768 such as C₁-C₆alkanoyloxy, C₀-C₃alkylaryloyloxy, particularly(optionally substituted) benzoyloxy or C₀-C₃alkylheterocycloyloxy,especially

This publication also describes alternative —W—R⁸ possibilities forexample C₁-C₆alkyl, such as ethyl, isopropyl, C₀-C₃alkylcarbocyclyl suchas cyclohexyl, 2,2-difluoroethyl, —C(═O)NRc, where Rc is C₁-C₆alkyl,C₀-C₃alkylcyclopropyl, C₀-C₃alkylaryl or C₀-C₃alkylheterocyclyl.

Currently preferred W functions include —S— and especially —O—.Convenient values for R⁸ in such embodiments include C₀-C₃alkylaryl, orC₀-C₃alkylheteroaryl either of which is optionally mono, di, or trisubstituted with R⁹, wherein;

R⁹ is C₁-C₆alkyl, C₁-C₆alkoxy, NO₂, OH, halo, trifluoromethyl, amino oramido (such as amino or amido optionally mono- or di-substituted withC₁-C₆alkyl), C₀-C₃alkylaryl, C₀-C₃alkylheteroaryl, carboxyl, aryl orheteroaryl being optionally substituted with R¹⁰; whereinR¹⁰ is C₁-C₆alkyl, C₃-C₇cycloalkyl, C₁-C₆alkoxy, amino (such as aminomono- or di-substituted with C₁-C₆alkyl), amido (such as C₁-C₅ alkylamide), sulfonylC₁-C₃alkyl, NO₂, OH, halo, trifluoromethyl, carboxyl, orheteroaryl.

Typically, the C₀-C₃ alkyl component of R⁸ as C₀-C₃alkylaryl, orC₀-C₃alkylheteroaryl is methyl and especially absent, ie C₀. The aryl orheteroaryl component s as extensively illustrated in the definitionsection below.

Preferred R⁹ include C₁-C₆ alkyl, C₁-C₆alkoxy, amino (such as di-(C₁-C₃alkyl)amino), amide (such as as —NHC(O)C₁-C₆alkyl or C(═O)NHC₁-C₃alkyl),aryl or heteroaryl, the aryl or heteroaryl moiety being optionallysubstituted with R¹⁰; wherein

R¹⁰ is C₁-C₆alkyl, C₃-C₇cycloalkyl, C₁-C₆alkoxy, amino (such as mono- ordi-C₁-C₃ alkylamino), amido (such as as —NHC(O)C₁-C₃yl orC(═O)NHC₁-C₆alkyl), halo, trifluoromethyl, or heteroaryl.

Preferred R¹⁰ include C₁-C₆alkyl, C₁-C₆alkoxy, amino, amido (such as as—NHC(O)C₁-C₆alkyl or C(═O)NHC₁-C₆alkyl) halo, or heteroaryl.

Particularly preferred R¹⁰ include methyl, ethyl, isopropyl, tertbutyl,methoxy, chloro, amino, amido (such as as —NHC(O)C₁-C₃alkyl orC(═O)NHC₁-C₆alkyl), or C₁-C₃alkyl thiazole.

Favoured embodiments of R₈ include 1-naphthylmethyl, 2-naphthylmethyl,benzyl, 1-naphthyl, 2-naphthyl, or quinolinyl, any opf which isunsubstituted, or mono- or disubstituted with R⁸ as defined, inparticular 1-naphthylmethyl, or quinolinyl unsubstituted, mono, ordisubstituted with R as defined.

A currently preferred R⁸ is:

wherein R^(9a) is C₁-C₆alkyl; C₁-C₆alkoxy; thioC₁-C₃alkyl; aminooptionally substituted with C₁-C₆alkyl; C₀-C₃alkylaryl; orC₀-C₃alkylheteroaryl, C₀-C₃alkylheterocyclyl, said aryl, heteroaryl orheterocycle being optionally substituted with R₁₀ wherein R¹⁰ isC₁-C₆alkyl, C₃-C₇cycloakyl, C₁-C₆alkoxy, amino, amido, heteroaryl orheterocyclyl; andR^(9b) is C₁-C₆alkyl, C₁-C₆alkoxy, amino, amido, NO₂, OH, halo,trifluoromethyl, carboxyl.

Convenient R^(9a) include aryl or heteroaryl, al optionally substitutedwith R¹⁰ as defined, especially where R^(9a) is selected from the groupconsisted of:

wherein R¹⁰ is H, C₁-C₆alkyl, or C₀-C₃alkyl-C₃-C₆cycloalkyl, amino (suchas amino mono- or di-substituted with C₁-C₆alkyl), amido (such as as—NHC(O)C₁-C₆alkyl or C(═O)NHC₁-C₆alkyl) heteroaryl or heterocyclyl.R^(9a) is conveniently phenyl and thus R⁸ is:

wherein R^(10a) is H, C₁-C₆alkyl; C₁-C₆alkoxy; or halo; and R^(9b) isC₁-C₆ alkyl, C₁-C₆-alkoxy, amino (such as C₁-C₃alkylamino), amido (suchas as —NHC(O)C₁-C₆alkyl or C(═O)NHC₁-C₃alkyl), NO₂, OH, halo,trifluoromethyl or carboxyl.

An alternative preferred R⁸ is:

wherein R^(10a) is H, C₁-C₆alkyl, or C₀-C₃alkyl-C₃-C₆cycloalkyl, amino(such as amino optionally mono- or di-substituted with C₁-C₆alkyl),amido (such as as —NHC(O)C₁-C₆alkyl or C(O)NHC₁-C₃alkyl orC(═O)N(C₁-C₃alkyl)₂), heteroaryl or heterocycyl; and R^(9b) is C₁-C₆alkyl, C₁-C₆-alkoxy, amino optionally mono- or di-substituted withC₁-C₆alkyl, amido (such as as —NHC(O)C₁-C₆alkyl or C(═O)NHC₁-C₃alkyl orC(═O)N(C₁-C₃alkyl)₃), NO₂, OH, halo, trifluoromethyl, or carboxyl.

In the immediately above described embodiments R^(9b) is convenientlyC₁-C₆-alkoxy, preferably methoxy.

A further R⁸ group, for example when W is an ether has the formula

where W is N or CH, r is 0 or 1, Ra′ is H, C₁-C₅ alkyl,C₀-C₃alkylcycloalkyl, C₁-C₆alkyloxy, hydroxy or amine and Rb′ is H,halo, C₁-C₆alkyl, C₀-C₃alkylcycloalkyl, C₁-C₆alkyloxy, C₁-C₆thioalkyl,cycloalkylC₀-C₃alkyloxy, C₁-C₃alkyloxyC₁-C₃alkyl, C₀-C₃alkylaryl orC₀-C₃alkylheterocyclyl A particularly preferred ether substituent is7-methoxy-2-phenyl-quinolin-4-yl oxy.

When W is a bond then R⁸ is preferably a substituted or unsubstitutedheterocyclic ring system as described in WO20041072243 or WO2004/113665.

Representative examples of R⁸ when W is a bond include the followingaromatics which may optionally be substituted: 1H-pyrrole, 1H-imidazole,1H-pyrazole, furan, thiophene, oxazole, thiazole, isoxazole,isothiazole, pyridine, pyridazine, pyrimidine, pyrazine, phthalazine,quinoxaline, quinazoline, quinoline, cinnoline,1H-pyrrolo[2,3]-b]pyridine, 1H-indole, 1H-benzoimidazole, 1H-indazole,7H-purine, benzothiazole, benzooxazole, 1H-imidazo[4, 5-c]pyridine,1H-imidazo[4,5-b]pyridine, 1, 3-dihydro-benzoimidazol-2-one, 1,3-dihydro-benzoimidazol-2-thione, 2, 3-dihydro-1H-indole,1,3-dihydro-indol-2-one, 1H-Indole-2,3-dione, 1,3-dihydro-benzoimidazole-2-one, 1H, 1H-pyrrolo [2, 3-c]pyridine,benzofuran, benzo[b]thiophene, benzo[d]isoxazole, benzo[d]isothiazol,1H-quinotin-2-one, 1H-quinolin-4-one, 1H-quinazol-4-one, 9H-carbazole,1H-quinazolin-2-one.

Additional representative examples of R⁸ when W is a bond, include thefollowing non-aromatics, which may be optionally substituted: aziridine,azetidine, pyrrolidine, 4,5-dihydro-1H-pyrazole, pyrazolidine,imidazolidin-2-one, imidazolidine-2-thione, pyrrolidin-2-one,pyrolidine-2,5-dione, piperidine-2,6-dione, piperidin-2-one,piperazine-2,6-done, piperazin-2-one, piperazine, morpholine,thiomorpholine-1,1-dioxide, pyrazolidin-3-one, imidazolidine-2,4-dione,piperidine, tetrahydrofuran, tetrahydropyran, [1,4]dioxane,1,2,3,6-tetrahydropyridine.

Preferred values for R⁸ when W is a bond, include tetrazole andderivatives thereof. The tetrazole moiety is linked to the cyclic P2scaffold and optionally substituted as shown below:

wherein Q* is selected from the group consisting of absent, —CH₂, —O—,—NH—, —N(R^(1*))—, —S—, —S(═O)₂— and —(C═O)—; Q* is selected from thegroup consisting of: absent, —CH₂— and —NH; Y* is selected from thegroup consisting of: H, C₁-C₆alkyl, C₀-C₃aryl, C₀-C₃heterocyclyl andR^(1*) is selected from the group consisting of: H, C₁-C₆alkyl,carbocyclyl, C₀-C₃aryl, C₀-C₃heterocyclyl.

Representative examples of substituted tetrazoles are as described intable 1 of WO2004/072243 and the structures following immediately after,or WO2004/113665.

Further preferred values for R⁸ when W is a bond, include triazole andderivatives thereof. The triazole moiety is linked to the cyclic P2scaffold and optionally substituted as shown below:

wherein X* and Y* are independently selected from the group consistingof: H, halogen, C₁-C₆alkyl, C₀-C₃carbocyclyl, —CH₂-amino,—CH₂-arylamino, —CH— diarylamino, —(C═O)-amino, —(C═O)-arylamino,—(C═O)-diarylamino, C₀-C₃aryl, C₀-C₃heterocyclyl or alternatively, X*and Y* taken together with the carbon atoms to which they are attached,form a cyclic moiety selected from the group consisting of aryl andheteroaryl.

Representative examples of substituted triazoles are as described intable 2 of WO2004/072243 and the structures following immediately after,or WO2004/113665.

Further preferred values for R⁸ when W is a bond, include pyridazinoneand derivatives thereof. The pyridazinone moiety is linked to the cyclicP2 scaffold and optionally substituted as shown below:

wherein X*, Y* and Z* are independently selected from the groupconsisting of: H, N₃, halogen, C₁-C₆alkyl, carbocyclyl, amino,C₀-C₃aryl, —S-aryl, —O-aryl, —NH-aryl, diarylamino, diheteroarylamino,C₀-C₃heterocycyl, —S-heteroaryl, —O-heteroaryl, NH-heteroaryl or,alternatively, X and Y or Y and Z taken together with the carbon atomsto which they are attached, form an aryl or heteroaryl cyclic moiety.

Representative examples of substituted pyridazinones are as described intable 3 of WO2004/072243 and the structures following immediately afteror WO2004/113865.

Preferred P3 groups, i.e. when m is 1 resemble natural or unnaturalamino adds, especially aliphatic amino adds, such as L-valyl, L-leucyl,L-isoleucyl or L-t-leucyl. Further preferred P3 groups, as shown in WO02/01898 include C₀-C₃alkylcycloalkylalanine, especiallycyclohexylalanine, optionally substituted with CO₂Rg, where Rg is H, isC₁-C₆alkyl, C₀-C₃alkylaryl, C₀-C₃alkylheterocyclyl, C₀-C₃alkylcycloalkylor amine; or N-acetylpiperidine or tetrahydropyran. Preferred R¹¹ groupsthus include C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl for exampleC₀-C₃alkylC₃-C₇cycloalkylyl, C₀-C₃alkylaryl or C₀-C₃alkylheteroaryl, anyof which is optionally substituted with hydroxy, halo, amino,C₁-C₆alkoxy, C₁-C₆thioalkyl, C(═O)OR¹⁴, carboxyl, (C₁-C₆alkoxy)carbonyl,aryl, heteroaryl or heterocyclyl, especially where the substituent ishydroxy or C(═O)OR¹⁴.

Particularly preferred R¹¹ include tert-butyl, iso-butyl, cyclohexyl,phenylethyl, 2,2-dimethyl-propyl, cyclohexylmethyl, phenylmethyl,2-pyridylmethyl, 4-hydroxy-phenylmethyl, or carboxylpropyl. The mostpreferred R¹¹ values are currently tert-butyl, Isobutyl, or cyclohexyl.

An embodiment of the invention include compounds wherein P4 is absent(ie n is 0) and wherein the P3 function lacks a carbonyl, ie U isabsent. Representative substructures include those of formula Ii below

wherein Rx and Ry are as defined above, preferably H,R^(11′) is C₁-C₆alkyl, preferably C₃-C₅ branched alkyl such as the sidechains of L-valyl, L-leucyl, L-Isoleucyl, L-t-leucyl; or C₀-C₂alkylC₃-C₇cycloalkyl such as cyclohexyl or cyclohexylmethyl;R^(16a) is —Rba, —S(═O)pRba, —C(═O)Rba;Rba is C₁-C₆ alkyl, C₀-C₃alkylheterocyclyl, C₀-C₃alkylcarbocyclyl.

Alternatively, compounds of partial structure Ii may be macrocyclisedbetween an appropriate value of R⁷ and one of Rx, Ry or R^(11′).

Representative embodiments of P3 groups which lack a carboxy function(ie variable U is absent) include those of formula VIia-VIid below:

where Ar is carbocyclyl or heterocycyl, especially aryl or heteroaryl,any of which is optionally substituted with R⁹. Although the partialstructures of Formulae VIia-VIid have been illustrated in the context ofa compound wherein k is 1 and q′ s 0, it will be apparent that suchconfigurations of Formula VIi apply also to other values of q′ and k.Similarly, although the partial structures of formulae VIic and VIidshow an R^(1l)group corresponding to leucine, it will be apparent thatthese configurations will be applicable to other R¹¹ groups, especiallythose resembling the side chains of natural or unnatural L-amino acids,for example t-butyl alanine/t.leucine.

R¹⁵ in those compounds of the invention wherein n is 1, is preferablyoptionally substituted C₁-C₆alkyl or C₀-C₃alkylcarbocyclyl for exampleC₀-C₃alkylC₃-C₇cycloalkyl, any of which may be optionally substituted.Preferred P4 groups are typically analogues of natural or unnaturalamino adds, especially aliphatic amino acids such as L-valyl, L-leucyl,L-isoleucyl, L-t-leucyl or L-cyclohexylalanine and thus favoured R¹⁵groups include cyclohexyl, cyclohexylmethyl, tert-butyl, iso-propyl, oriso-butyl.

Preferred G values include —NRy-, especially wherein Ry is methyl orpreferably H, or hydrazine.

A further preferred G value is O thereby defining an ester with thecarbonyl of P4 (if present) or the carbonyl of P3 (if present) or anether in the case of variants wherein group U is absent. Conventionalpharmaceutically acceptable ethers or esters capping groups for R¹include C₁-C₆alkyl (especially methyl or t-butyl),C₀-C₃alkylheterocyclyl (especially pyridyl, benzimidazolyl, piperidyl,morpholinyl, piperazinyl) or C₀-C₃alkylcarbocyclyl (especially phenyl,benzyl, indanyl) any of which is optionally substituted with hydroxy,halo, amino, or C₁-C₆alkoxy.

Favoured compounds of the invention can comprise a hydrazinefunctionality, for example where T is —NRd- and m is 1; with n beingzero or 1. Alternatively, especially where m is zero, G can be —NRjNRj-such as —NHNH—. Compounds will generally not comprise a hydrazine atboth G and T. Preferred hydrazines within Formula VI, wherein m and nare zero include compounds of the partial structures VIja-VIjb below:

R¹⁶ in formulae VIja and VIjb can be regarded as an alkyl (orC₁-C₃alkylheterocyclyl or C₁-C₃alkylcarbocyclyl) wherein the first alkylcarbon is substituted with an oxo group to define the keto function andR^(16′) is the remainder of the alkyl, alkylheterocyclyl oralkylcarbocyclyl moiety. Formula VIjb depicts a variant where R¹⁶ is amethylene group whose carbon is substituted with an oxo substituent andalso with —ORb, where Rb is as defined above, typically, C₁-C₆alkyl,such as t-butyl, C₀-C₃alkylheterocyclyl such as pyridyl, orC₀-C₃alkylcarbocyclyl, such as benzyl or phenyl, any of which isoptionally substituted as defined above. Compounds of partial structuresVIja and VIjb can be linear molecules as shown (both Rj are H), orpreferably one of the depicted Rj groups can be macrocyclised via J toan appropriate R⁷ group.

Alternative hydrazines of Formula VI where m is 1 include those ofpartial structures VIjc and VIjd below

where G, R¹⁶, R¹⁶, Rx, Rd, Rq, Rz, and Ru are as defined for formula VIabove. Compounds of partial structures VIjc and VIjd can be linearmolecules as shown (both Rx and Rd are H), or preferably one of thedepicted Rx or Rd groups can be macrocyclised via J to an appropriate R⁷group.

Although formulae VIja-VIjd are depicted with a five membered carbocylicring as P2 scaffold, it will be apparent that this aspect of theinvention is equally adapted to other configurations of q′ and k.Favoured embodiments within formula VIja-VIjd include those wherein Rqand Rz are H, or those wherein Rz is an olefinic bond and Rq isC₁-C₃alkyl.

Alternative hydrazine-like configuration are found when G is amino, andm and n are 0, and R¹⁶ is an N-linked unsaturated heterocycle as definedbelow, for example pyridyl or pyrimidyl or a saturated heterocycle asdefined below, such as piperazinyl, piperidinyl and especiallymorpholinyl. Examples of such embodiments include those of the formulaeVIjc and VIjd:

Compounds of partial structures VIjc and VIjd can be linear molecules asshown or preferably Rx can be macrocyclised via J to an appropriate R⁷group. Although these partial structures are depicted with a fivemembered ring as the P2 scaffold, it will be readily apparent that thisconfiguration extends to other values of q′ and k. Similarly theseconfigurations will be applicable to other N-linked heterocycles as R¹⁶.

Returning now to Formulae VI in general, favoured R¹⁶ groups for thecompounds of the invention include 2-indanol, indanyl,2-hydroxy-1-phenyl-ethyl, 2-thiophenemethyl, cyclohexylmethyl,2,3-methylenedioxybenzyl, cyclohexyl, phenyl, benzyl, 2-pyridylmethyl,cyclobutyl, iso-butyl, n-propyl, methyl, or 4-methoxyphenylethyl.

Currently preferred R¹⁶ groups include 2-indanol, indan,2-hydroxy-1-phenyl-ethyl, 2-thiophenemethyl, 2,3-methylenedioxybenzyl,or cyclohexylmethyl.

Unnatural amino adds include L-amino adds wherein the side chain is notone of the 20 naturally occurring amino acids. Examples of non-naturalamino adds include L-beta-methylsulfonylmethylalanine,L-cyclohexylalanine, L-tertiary-leucine, L-norleucine, L-norvaline,L-ornithine, L-sarcosine, L-citurline, L-homophenylalanine,L-homoserine, L-beta-(1-napthyl)alanine, L-beta-(2-napthyl)alanine etc.Non natural amino acids also include the D-amino acids corresponding tothe 20 natural amino adds and D-amino acids bearing other side chains,such as those listed above.

‘C₁-C₆alkyl’ (also abbreviated as C₁-C₆alk, or used in compoundexpressions such as C₁-C₆alkyloxy etc) as applied herein is meant toinclude straight and branched chain aliphatic carbon chains such asmethyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl, pentyl,isopentyl, hexyl, heptyl and any simple isomers thereof. The alkyl groupmay have an unsaturated bond. Additionally, any C atom in C₁-C₆alkyl mayoptionally be substituted by one, two or where valency permits threehalogens and/or substituted or the alkylene chain interrupted by aheteroatom S, O, NH. If the heteroatom is located at a chain terminusthen it is appropriately substituted with one or 2 hydrogen atoms.C₁-C₄alkyl and C₁-C₆alkyl have the corresponding meaning to C₁-C₆alkyladjusted as necessary for the carbon number.

‘C₁-C₃alkyl’ as applied herein includes methyl, ethyl, propyl,isopropyl, cyclopropyl, any of which may be optionally substituted orheteroatom interrupted as described in the paragraph above or in thecase of C₂ or C₃, bear an unsaturated bond such as CH₂═CH.

“C₁-C₃ alkylene” as applied herein describes a divalent C1-C3alkyldiylmoiety, Including propylene, ethylene and especially methylene. Thetypically longer alkylene chains for J may comprise 1 to 3 unsaturationsand/or interruptions with heteroatoms as defined above.

‘Amino’ includes NH₂, NHC₁-C₆alkyl or N(C₁-C₆-alkyl), especially C₁-C₃alkyl variants

‘Amido’ includes C(═O)NH₂ and alkylamido such as C(═O)NHC₁-C₆alkyl,C(═O)N(C₁-C₆alkyl)₂ especially C(═O)NHC₁-C₃alkyl, C(O)N(C₁-C₃alkyl)₂ or—NH(C═O)C₁-C₆alkyl, for example —NHC(═O)CHC(CH₃)₃, including—NH(C═O)C₁-C₃alkyl.

‘Halo’ or halogen as applied herein is meant to include F, Cl, Br, I,particularly chloro and preferably fluoro.

‘C₀-C₃alkylaryl’ as applied herein is meant to include an aryl moietysuch as a phenyl, naphthyl or phenyl fused to a C₃-C₇cycloalkyl forexample indanyl, which amyl is directly bonded (i.e. C₀) or through anintermediate methyl, ethyl, propyl, or isopropyl group as defined forC₁-C₃alkylene above. Unless otherwise indicated the aryl and/or itsfused cycloalkyl moiety is optionally substituted with 1-3 substituentsselected from halo, hydroxy, nitro, cyano, carboxy, C₁-C₆alkyl,C₁-C₆alkoxy, C₁-C₆alkoxyC₁-C₆alkyl, C₁-C₆alkanoyl, amino, azido, oxo,mercapto, nitro C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl. “Aryl”has the corresponding meaning, i.e. where the C₀-C₃alkyl linkage isabsent.

‘C₀-C₃alkylC₃-C₇cycloalkyl’ as applied herein is meant to include aC₃-C₇cycloalkyl group such as cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl or cycloheptyl, which cycloakyl is directly bonded (i.e.C₀alkyl) or through an intermediate methyl, ethyl or proyl group asdefined for C₁-C₃alkylene above. The cycloalkyl group may contain anunsaturated bond. Unless otherwise indicated the cycloalkyl moiety isoptionally substituted with 1-3 substituents selected from halo,hydroxy, nitro, cyano, carboxy, C₁-C₆alkyl, C₁-C₆alkoxy,C₁-C₆alkoxyC₁-C₆alkyl, C₁-C₆alkanoyl, amino, azido, oxo, mercapto, nitroC₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl.

‘C₀-C₃alkylcarbocyclyl’ as applied herein is meant to includeC₀-C₃alkylaryl and C₀-C₃alkylC₃-C₇cycloalkyl. Unless otherwise indicatedthe aryl or cycloalkyl group is optionally substituted with 1-3substituents selected from halo, hydroxy, nitro, cyano, carboxy,C₁-C₆alkyl, C₁-C₆alkoxy, C₁-C₆alkoxyC₁-C₆alkyl, C₁-C₆alkanoyl, amino,azido, oxo, mercapto, nitro, C₀-C₃alkylcarbocyclyl and/orC₀-C₃alkylheterocyclyl. “Carbocyclyl” has the corresponding meaning,i.e. where the C₀-C₃alkyl linkage is absent

‘C₀-C₃alkylheterocycylyl’ as applied herein is meant to include amonocyclic, saturated or unsaturated, heteroatom-containing ring such aspiperidinyl, morpholinyl, piperazinyl, pyrazolyl, imidazolyl, oxazolyl,isoxazolyl, thiazinolyl, isothiazinolyl, thiazolyl, oxadiazolyl,1,2,3-triazolyl, 1,2,4-triazolyl, tetrazolyl, furanyl, thienyl, pyridyl,pyrimidyl, pyridazinyl, pyrazolyl, or any of such groups fused to aphenyl ring, such as quinolinyl, benzimidazolyl, benzoxazolyl,benzisoxazolyl, benzothiazinolyl, benzisothiazinolyl, benzothiazolyl,benzoxadiazolyl, benzo-1,2,3-triazolyl, benzo-1,2,4-triazolyl,benzotetrazolyl, benzofuranyl, benzothienyl, benzopyridyl,benzopyrimidyl, benzopyridazinyl, benzopyrazolyl etc, which ring isbonded directly i.e. (C₀), or through an intermediate methyl, ethyl,propyl, or isopropyl group as defined for C₁-C₃alkylene above. Any suchnon-saturated rings having an aromatic character may be referred to asheteroaryl herein. Unless otherwise indicated the hetero ring and/or itsfused phenyl moiety is optionally substituted with 1-3 substituentsselected from halo, hydroxy, nitro, cyano, carboxy, C₁-C₆alkyl,C₁-C₆alkoxy, C₁-C₆alkoxyC₁-C₆alkyl, C₁-C₆alkanoyl, amino, azido, oxo,mercapto, nitro, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl.“Heterocycly” and “Heteroaryl” have the corresponding meaning, i.e.where the C₀-C₃alkyl linkage is absent.

Typically heterocycyl and carbocyclyl moieties within the scope of theabove definitions are thus a monocyclic ring with 5 or especially 6 ringatoms, or a bicyclic ring structure comprising a 6 membered ring fusedto a 4, 5 or 6 membered ring.

Typical such groups include C₃-C₈cycloalkyl, phenyl, benzyl,tetrahydronaphthyl, Indenyl, indanyl, heterocyclyl such as fromazepanyl, azocanyl, pyrrolidinyl, piperidinyl, morpholinyl,thiomorpholinyl, piperazinyl, indolinyl, pyranyl, tetrahydropyranyl,tetrahydrothiopyranyl, thiopyranyl, furanyl, tetrahydrofuranyl, thienyl,pyrrolyl, oxazolyl, isoxazolyl, thiazolyl, imidazolyl, pyridinyl,pyrimidinyl, pyrazinyl, pyridazinyl, tetrazolyl, pyrazolyl, indolyl,benzofuranyl, benzothienyl, benzimidazolyl, benzthiazolyl, benzoxazolyl,benzisoxazolyl, quinolinyl, tetrahydroquinolinyl, isoquinolinyl,tetrahydroisoquinolinyl, quinazolinyl, tetrahydroquinazolinyl andquinoxalinyl, any of which may be optionally substituted as definedherein.

The saturated heterocycle moiety thus includes radicals such aspyrrolinyl, pyrrolidinyl, pyrazolinyl, pyrazolidinyl, piperidinyl,morpholinyl, thiomorpholinyl, pyranyl, thiopyranyl, piperazinyl,indolinyl, azetidinyl, tetrahydropyranyl, tetrahydrothiopyranyl,tetrahydrofuranyl, hexahydropyrimidinyl, hexahydropyridazinyl,1,4,5,6-tetrahydropyrimidinylamine, dihydro-oxazolyl,1,2-thiazinanyl-1,1-dioxide, 1,2,6-thiadiazinanyl-1,1-dioxide,isothiazolidinyl-1,1-dioxide and imidazolidinyl-2,4-dione, whereas theunsaturated heterocycle include radicals with an aromatic character suchas furanyl, thenyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl,pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, tetrazolyl,thiadiazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl,indolizinyl, indolyl, isoindolyl. In each case the heterocycle may becondensed with a phenyl ring to form a bicyclic ring system.

Synthesis

Synthesis of the compounds of the present invention can be performed bydifferent chemical strategies in solution or solid phase or acombination of both. The suitably protected individual building blockscan first be prepared and subsequently coupled together i.e.P2+P1→P2-P1. Alternatively, precursors of the building blocks can becoupled together and modified at a later stage of the synthesis of theinhibitor sequence. Further building blocks, precursors of buildingblocks or prefabricated bigger fragments of the desired structure, canthen be coupled to the growing chain, e.g.R¹⁶-G-P3+C(═O)—P2-P1→R¹⁶-G-P3-C(═O)—P2-P1 orR¹⁶-G-P4-P3+C(═O)—P2-P1→R1-G-P4-P3-C(═O)—P2-P1.

Coupling between two amino acids, an amino acid and a peptide, or twopeptide fragments can be carried out using standard coupling proceduressuch as the azide method, mixed carbonic-carboxylic acid anhydride(Isobutyl chloroformate) method, carbodiimide (dicyclohexylcarbodiimide,diisopropylcarbodiimide, or water-soluble carbodiimide) method, activeester (pnitrophenyl ester, N-hydroxysuccinic imido ester) method,Woodward reagent K-method, carbonyldiimidazole method, phosphorusreagents or oxidation-reduction methods. Some of these methods(especially the carbodimide method) can be enhanced by adding1-hydroxybenzotriazole or 4-DMAP. These coupling reactions can beperformed in either solution (liquid phase) or solid phase.

More explicitly, the coupling step involves the dehydrative coupling ofa free carboxyl of one reactant with the free amino group of the otherreactant in the present of a coupling agent to form a linking amidebond. Descriptions of such coupling agents are found in generaltextbooks on peptide chemistry, for example, M. Bodanszky, “PeptideChemistry”, 2nd rev ed., Springer-Verlag, Berlin, Germany, (1993)hereafter simply referred to as Bodanszky, the contents of which arehereby incorporated by reference. Examples of suitable coupling agentsare N,N′-dicyclohexylcarbodiimide, 1-hydroxybenzotriazole in thepresence of N,N′-dicyclohexylcarbodiimide orN-ethyl-N′-[(3dimethylamino) propyl] carbodiimide. A practical anduseful coupling agent is the commercially available(benzotriazol-1-yloxy) tris-(dimethylamino) phosphoniumhexafluorophosphate, either by itself or in the present of1-hydroxybenzotriazole or 4-DMAP. Another practical and useful couplingagent is commercially available 2-(1H-benzotriazol-1-yl)-N, N,N′,N′-tetramethyluronium tetrafluoroborate. Still another practical anduseful coupling agent is commercially available0-(7-azabenzotrizol-1-yl)-N, N,N′, N′-tetramethyluroniumhexafluorophosphate.

The coupling reaction is conducted in an inert solvent, e.g.dichloromethane, acetonitrile or dimethylformamide. An excess of atertiary amine, e.g. diisopropylethylamine, N-methylmorpholine,N-methylpyrrolidine or 4-DMAP is added to maintain the reaction mixtureat a pH of about 8. The reaction temperature usually ranges between 0°C. and 50° C. and the reaction time usually ranges between 15 min and 24h.

The functional groups of the constituent amino acids generally must beprotected during the coupling reactions to avoid formation of undesiredbonds. The protecting groups that can be used are listed in Greene,“Protective Groups in Organic Chemistry”, John Wiley & Sons, New York(1981) and “The Peptides: Analysis, Synthesis, Biology”, Vol. 3,Academic Press, New York (1981), hereafter referred to simply as Greene,the disclosures of which are hereby incorporated by reference.

The α-carboxyl group of the C-terminal residue is usually protected asan ester that can be cleaved to give the carboxylic acid. Protectinggroups that can be used include 1) alkyl esters such as methyl,trimethylsilyl and t.butyl, 2) aralkyl esters such as benzyl andsubstituted benzyl, or 3) esters that can be cleaved by mild base ormild reductive means such as trichloroethyl and phenacyl esters.

The α-amino group of each amino acid to be coupled is typicallyprotected. Any protecting group known in the art can be used. Examplesof such groups include: 1) acyl groups such as formyl, trifluoroacetyl,phthalyl, and p-toluenesulfonyl; 2) aromatic carbamate groups such asbenzyloxycarbonyl (Cbz or Z) and substituted bensyloxycarbonyls, and9-fluorenylmethyloxycarbonyl (Fmoc); 3) aliphatic carbamate groups suchas tertbutyloxycarbonyl (Boc), ethoxycarbonyl,diisopropylmethoxycarbonyl, and allyloxycarbonyl; 4) cyclic alkylcarbamate groups such as cyclopentyloxycarbonyl and adamantyloxycarbonyl5) alkyl groups such as triphenylmethyl and benzyl; 6) trialkylsilylsuch as trimethylsilyl; and 7) thiol containing groups suchasphenylthiocarbonyl anddithiasuccinoyl. The preferred α-aminoprotecting group is either Boc or Fmoc. Many amino acid derivativessuitably protected for peptide synthesis are commercially available.

The α-amino protecting group is cleaved prior to the next coupling step.When the Boc group is used, the methods of choice are trifluoroaceticacid, neat or in dichloromethane, or HCl in dioxane or in ethyl acetate.The resulting ammonium salt is then neutralized either prior to thecoupling or in situ with basic solutions such as aqueous buffers, ortertiary amines in dichloromethane or acetonitrile or dimethylformamide.When the Fmoc group is used, the reagents of choice are piperidine orsubstituted piperidine in dimethylformamide, but any secondary amine canbe used. The deprotection is carried out at a temperature between 0° C.and room temperature usually 20-22° C.

Any of the natural or non-natural amino acids having side chainfunctionalities will typically be protected during the preparation ofthe peptide using any of the above described groups. Those skilled inthe art will appreciate that the selection and use of appropriateprotecting groups for these side chain functionalities depend upon theamino acid and presence of other protecting groups in the peptide. Inthe selection of such protecting groups it is desirable that the groupis not removed during the deprotection and coupling of the α-aminogroup.

For example, when Boc is used as the α-amino protecting group, thefollowing side chain protecting groups are suitable: p-toluenesulfonyl(tosyl) moieties can be used to protect the amino side chain of aminoacids such as Lys and Arg; acetamidomethyl, benzyl (Bn), ortert-butylsulfonyl moities can be used to protect the sulfide containingside chain of cysteine; benzyl (Bn) ethers can be used to protect thehydroxy containing side chains of serine, threonine or hydroxyproline;and benzyl esters can be used to protect the carboxy containing sidechains of aspartic acid and glutamic acid.

When Fmoc is chosen for the α-amine protection, usually tert. butylbased protecting groups are acceptable. For instance, Boc can be usedfor lysine and arginine, tert.butyl ether for seine, threonine andhydroxyproline, and tert-butyl ester for aspartic acid and glutamicacid. Triphenylmethyl (Trityl) moiety can be used to protect the sulfidecontaining side chain of cysteine.

Once the inhibitor sequence is completed any protecting groups areremoved in whatever manner is dictated by the choice of protectinggroups. These procedures are well known to those sidled in the art.

Introduction of the P2 Substituent

The R⁸ group can be coupled to the P2 scaffold at any convenient stageof the synthesis of compounds according to the present invention. Oneapproach is to first couple the R⁸ group to the P2 scaffold andsubsequently add the other desired building blocks, i.e. P1 andoptionally P3 and P4. Another approach is to couple the P1, P2 and ifpresent P3 and P4 moieties using an unsubstituted P2 scaffold and addthe R⁸ group afterwards.

Compounds wherein W is O and R⁸ is alkyl, C₀-C₃alkylcarbocycylyl,C₀-C₃alkylheterocycylyl can be prepared according to the proceduredescribed by E. M. Smith et al. (J. Med. Chem. (1988), 31, 875-885), asdepicted in Scheme 1, which illustrates the technique with a saturatedP2 scaffold wherein q′ is 0 and k is 1.

Treatment of a compound containing an unsubstituted P2 scaffold (1a),which ca be prepared as described herein below with a base such assodium hydride or potassium t.butoxide in a solvent likedimethylformamide followed by reaction of the resulting alkoxide with analkylating agent, R⁸—X, wherein X is a suitable leaving group such as ahalide, mesylate, triflate or tosylate, provides the desired substitutedderivative (1b).

Alternatively, if X is OH or SH, the P2 substituent can be introducedvia a Mitsunobu reaction by reacting the hydroxy group of compound 1awith the desired alcohol or thiol in the presence of triphenylphosphineand an activating agent like diethyl azodicarboxylate (DEAD),diisopropyl azodicarboxylate (DIAD) or the like. (Mitsunobu, 1981,Synthesis, January, 1-28; Rano et al., Tetrahedron Lett., 1995, 36, 22,3779-3792; Krchnak et al., Tetrahedron Lett., 1995, 36, 5, 6193-6196;Richter et al., Tetrahedron Lett., 1994, 35, 27, 4705-4706).

Alcohol (1a) can alternatively be treated with phosgene thus providingthe corresponding chloroformate which upon reaction with an amine,R⁸NH₂, in the presence of a base like sodium hydrogen carbonate ortriethylamine, provides carbamates i.e. W is —OC(═O)NH—, whereasreaction of alcohol (1a) with an acylating agent, R⁸—CO—X, like an acidanhydride or add halide for instance the acid chloride, to provideesters, i.e. W is —OC(═O)—.

Various alcohols R⁸—OH, and alkylating agents R⁸—X are described in WO00/09543 and WO00/59929. An example of the synthesis wherein R⁸ is asubstituted quinoline derivative is shown in Scheme 2.

Friedel-Craft acylation of a suitable substituted aniline (2a),available either commercially or in the literature, using an acylatingagent like acetyl chloride or the like in the presence of borontrichloride and aluminium trichloride in a solvent lice dichloromethaneprovides (2b). Coupling of (2b) to a heterocyclic carboxylic add (2c)under basic conditions, such as in pyridine, in the presence of anactivating agent for the carboxylate group, for instance POCl₃, followedby ring closure and dehydration under basic conditions like potassiumtert-butoxide in tert-butanol provides quinoline derivative (2e).Quinoline derivative (2e) can be coupled in a Mitsunobu reaction to analcohol as described above, or the hydroxy group can be displaced by asuitable leaving group such as a halide like chloride, bromide oriodide, by treatment of quinoline (2e) with an appropriate halogenatingagent for example phosphoryl chloride or the like.

A variety of carboxylic adds with the general structure (2c) can be usedin Scheme 2. These adds are available either commercially or in theliterature. An example of the preparation of2-(substituted)-amino-carboxy-aminothiazole derivatives, following theprocedure by Berdikhina et al. Chem. Heterocycl. Compd. (Eng. Transl.)(1991), 427-433, is shown in scheme 3 below.

Thiourea (3c) with different alkyl substituents R′ can be formed byreaction of the appropriate amine (3a) with tert-butylisothiocyanate inthe presence of a base like diisopropylethylamine in a solvent likedichloromethane followed by removal of the tert-butyl group under acidicconditions. Subsequent condensation of thiourea derivative (3c) with3-bromopyruvic acid provides the add (3d).

P2 building blocks wherein the R⁸ substituent is attached via an amine,amide, urea or sulphonamide, can be prepared from amino substitutedcarbocycles achieved for example by transforming the hydroxy group ofthe corresponding hydroxy derivative into an azide group for example bytransforming the hydroxy group into a suitable leaving group such as amesylate or halogen like chloride, followed by substitution of theleaving group with azide or by the use of an azide transfer agent likediphenylphosphoryl azide (DPPA). Reduction of the azide by catalytichydrogenation or any other suitable reduction method provides the amine.The amino derivative can be reacted in a displacement reaction with analkylating agent of the general formula R⁸—X wherein R⁸ and X are asdescribed for scheme 1, to form P2 building blocks for use in thepreparation of compounds of general formula VI, wherein W is —NH—.Reaction of the amino substituted carbocycle with an acid of the generalformula R⁸—COOH under standard amide coupling conditions providescompounds wherein the R⁸ substituent is linked via an amide bond,whereas reaction of the amino substituted carbocycle with an appropriatederivative of sulphonic acid, R⁸—S(O)₂—X where X is a leaving group forexample chloride, in the presence of a base, provides sulphonamides.Compounds wherein the linkage between the cyclic scaffold and the R⁸substituent is constituted of a urea group can for example be achievedby treatment of amino substituted carbocycle with phosgene to afford thecorresponding chlorocarbamate followed by reaction with the desiredamine. Alternatively, the amino substituted carbocycle can be reactedwith the carbamoyl chloride or isocyanate of the desired R⁸ substituentfor the formation of the urea linkage. It will be apparent thatcorresponding reactions will be available for P2 groups with other ringsizes and substitution pattern.

Compounds wherein a heterocyclic R⁸ group is attached directly to thecyclic P2 scaffold i.e. W is a bond in general formula VI, can beprepared for example by using a replacement reaction wherein a suitableleaving group such as halide a or a mesylate or the like on the P2scaffold is replaced by the desired R⁸ group such as a heterocyclicgroup. Alternatively the R^(e) group can be introduced by way of aMitsunobu reaction wherein the hydroxy group of the P2 precursor isreacted with a nitrogen atom in the heterocyclic R⁸ group.

Compounds wherein a tetrazole derivative is attached to one of its ringcarbons are conveniently prepared by building up the tetrazole moietydirectly on the P2 precursor. This can be achieved for instance bytransforming the hydroxy group of the P2 precursor into a cyano groupfollowed by reaction with an azide reagent like sodium azide. Triazolederivatives can also be built up directly on the P2 precursor forexample by transforming the hydroxy group of the P2 precursor into anazide group followed by a 3+2 cycloaddition reaction of the affordedazide and a suitable alkyne derivative.

Structurally diverse tetrazoles for use in the above describedsubstitution or Mitsunobu reactions can be prepared by reactingcommercially available nitrile compounds with sodium azide. Triazolederivatives can be prepared by reaction of an alkyne compound andtrimethylsilyl azide. Useful alkyne compounds are available eithercommercially or they can be prepared for instance according to theSonogashira reaction i.e. reaction of a primary alkyne, an aryl halideand triethylamine in the presence of PdCl₂(PPh)₃ and CuI as describedfor example in A. Elangovan, Y.-H. Wang, T.-I. Ho, Org. Let., 2003, 5,1841-1844. The heterocyclic substituent can also be modified whenattached to the P2 building block either before or after coupling of theP2 building block to the other building blocks.

These methods and further alternatives for the preparation of compoundswherein W is a bond and R⁸ is an optionally substituted heterocycle areextensively described in WO2004/072243.

Compounds having an alternative ring size and/or position of the W—R⁸substituent of the carbocyclic derivative in scheme 1 may also be usedin the preparation of compounds according to the present invention.

Synthesis and Introduction of P1 Building Blocks.

The amino acids used in the preparation of P1 fragments are availableeither commercially or in the literature, see for example WO 00/09543and WO00/59929 from Boehringer-Ingelhelm or US2004/0048802 from BMS.

Scheme 4 shows an example of the preparation of a sulphonamidederivative to be used as a P1 building block, and the subsequentcoupling to a P2 building block.

The sulphonamide group can be introduced on a suitably protected aminoacid (4a) by treatment of the amino acid with a coupling agent, forexample N,N′-carbonyldiimidazole (CDI) or the like, in a solvent likeTHF followed by reaction with the desired sulphonamide (6b) in thepresence of a strong base such as 1,8-diazabicyclo[5.4.0]undec-7-ene(DBU). Alternatively the amino acid can be treated with the desiredsulphonamide (4b) in the presence of a base like diisopropyl ethylaminefollowed by treatment with a coupling agent like PyBOP® to effect theintroduction of the sulphonamide group. Removal of the amino protectinggroup by standard methods and subsequent coupling to a P2 buildingblock, prepared as described below, using standard methods for amidebond formation, like using a coupling agent such asO-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (HATU) in the presence of a base such asdiisopropylamine in a solvent like dimethylformamide, gives 4e.Alternatively, the sulphonamide group can be introduced at a later stageof the synthesis, for example as the last step. In this case an aminoacid with the reversed protection pattern, i.e. with an unprotectedamino function and protected acid function, is coupled the acid functionof the P2 building block using standard peptide coupling conditions forexample as described above. Removal of the acid protection group, usingthe appropriate conditions for the present protection group, followed bycoupling of the sulphonamide as described above then yields compound 4e.

P1 building blocks for the preparation of compounds according to generalformula VI wherein A is an ester or an amide can be prepared by reactingamino acid (4a) with the appropriate amine or alcohol respectively understandard conditions for amide or ester formation. Compounds according togeneral formula I wherein A is CR⁴R^(4′) can be prepared by coupling ofthe appropriate P1 building block to the P2 building block as describedin Oscarsson et al Bioorg Med Chem 2003 11(13) 2955-2963 andPCT/EP03/10595 filed 23 Sep. 2003, the contents of which areincorporated by reference.

Compounds comprising an azapeptide P1 residue, i.e. M is NRu in generalformula VI can be prepared by using a suitable P1 aza-amino acyl moietyin the coupling to the P2 fragment. The preparation of aza-amino acylmoieties is described by M. D. Bailey at al. In J. Med. Chem., 47,(2004), 3788-3799, and an example is shown in scheme 5.

Incorporation of the appropriate N-linked side chain, Ru, oncommercially available tert-butylhydrazine can be performed for exampleby a reductive amination reaction with the appropriate aldehyde orketone as described in scheme 19 below which produces the N-alkylatedcarbazate (5a). Condensation of 5a with a desired chloroformate in thepresence of a base like triethylamine or diisopropylethylamine in asolvent like THF provides 5b. The R1′ moiety can then optionally beremoved using the appropriate conditions depending on the specific R1′,such as catalytic hydrogenation for R1′ being benzyl, which gives thecorresponding acids. Subsequent reaction of the afforded acid with adesired sulphonamide derivative as described in scheme 4 yieldssulphonamide capped building blocks. Alternatively, reaction ofcarbazate 5a with an isocyanate, R3-N═C═O, provides building blocks forthe preparation of compounds according to general formula VI wherein Mis NRu and A is CONHR³.

Synthesis of Capped P3 and P4-P3 Building Blocks

The building blocks R¹⁶-G-P3 and R¹⁶-G-P4-P3 can be prepared asgenerally depicted in scheme 6.

A suitable N-protected amino acid (6a) can be coupled with an aminocapping group (R¹⁶—NHRy) using standard peptide coupling conditions likewith coupling agents such as HATU, DCC, HOBt or the like in the presenceof a base such as DIEA or DMAP in a solvent like dichloromethane,chloroform or dimethylformamide or a mixture thereof and ester formationconditions like providing amides i.e. G is NHRy (6b). Alternatively,reaction of amino acid (6a) with a compound of general formula R¹⁶—Xwhere R¹⁶ is as defined above and X is a leaving group such as a halide,in the presence of a base like cesium carbonate or silver (I) oxideprovides esters, i.e. G is O (6b). On the other hand, amino acid (6a)can be coupled to a second, suitably O-protected, amino acid (6d) usingstandard peptide coupling conditions as described above, providing (6e).Displacement of the ester group with a suitable capping group (6b)provides fragment (6f) useful for the preparation of compounds accordingto the present invention wherein m and n are 1.

When G is N-Ry, the capped P3 or P2 building block can also be preparedon solid support as exemplified in Scheme 7.

An appropriate N-protected, for example Boc protected, amino acid (7a)can be immobilized on a solid support, here exemplified by Agronautresin PS-TFP, by reacting the amino acid with the desired solid supportin the presence of coupling reagent like N,N′-diisopropylcarbodiimideand a base like DMAP in a solvent like dichloromethane anddimethylformamide. The immobilized amino acid (7b) can then be cleavedfrom the support with a suitable capping group (7c) thus givingfragments (7d) useful for the preparation of compounds according to thepresent invention wherein m or n is 1. Optionally the amino protectinggroup can be removed followed by coupling of an appropriate amino acidusing standard methods thus providing fragments useful for thepreparation of compounds according to the present invention wherein mand n are 1.

Preparation and Incorporation of P2 Building Blocks

A typical route to compounds containing a 5 membered saturated P2scaffold is shown in Scheme 8.

The cyclic scaffold (8b) can be prepared, for example, from3,4-bis(methoxycarbonyl)cyclopentanone (8a), as described by Rosenquistet al. in Acts Chem. Scand. 46 (1992) 1127-1129 by reduction of the ketogroup with a reduction agent like sodium borohydride in a solvent likemethanol followed by hydrolysis of the esters and finally ring closurein acetic anhydride in the presence of pyridine. The provided bicyclicacid (8b) can then be coupled to the amine function of the desired P3fragment (8c), P3-P4 fragment or capping group R¹⁶—NHRy, usingconventional peptide coupling conditions like with HATU and diisopropylamine in a solvent like dimethyl formamide, giving (8d). Lactone openingof (8d) with for example lithium hydroxide provides the acid whichsubsequently can be coupled to the amino group of a P1 building block ora precursor of a desired P1 fragment (8e), using conventional peptidecoupling conditions. The R⁸-substituent of the carbocycle can beintroduced for example by a Mitsunobu reaction with the appropriatealcohol as described above or by any other suitable method previouslydescribed. When R⁷, R^(7′) and A′ contains functional groups, these areoptionally suitably protected by methods recognized by persons sidled inthe art, se for example Bodanzky or Greene cited above.

Scheme 9 shows an alternative route towards compounds of formula VIcontaining a saturated P2 scaffold, where the building blocks areintroduced in the reversed order, i.e. the P1 fragment is introducedbefore the capping group, P3 or P3-P4 building block.

Protection of the add group of (9a) for example as the tert-butyl esterby treatment with dl-tert-butyl dicarbonate in the presence of a baselike dimethylaminopyridine and triethylamine in a solvent likedichloromethane provides ester (9b). Lactone opening and coupling of aP1 building block (9c) as described in scheme 13 or directly by theamine group of the P1 fragment provides (9d). Introduction ofR⁸-substituent as described above followed by removal of the acidprotection group by subjecting the ester to acidic conditions liketrifluoroacetic acid and triethylsilane in a solvent like methylenechloride and finally coupling of the P3 building block (9e), P3-P4building block or capping group R¹⁶—NHRy, as described above provides(9f). When R⁷, R^(7′) and A′ contain functional groups, these areoptionally suitably protected by methods recognized by persons skilledin the art, see for example Bodanzky or Greene cited above.

An unsaturated P2 scaffold to be used in the preparation of compounds offormula VI can be prepared as illustrated with cyclopentene below.

The cyclopentene scaffold is typically prepared as described in scheme10.

A bromination-elimination reaction of3,4-bis(methoxycarbonyl)cyclopentanone (10a) as described by Dolby etal. in J. Org. Chem. 36 (1971) 1277-1285 followed by reduction of theketo functionality with a reduction agent like sodium borohydrideprovides the unsaturated hydroxy compound (10b). Selective esterhydrolysis using for example lithium hydroxide in a solvent like amixture of dioxane and water provides hydroxy substituted monoesterderivative (10c).

An unsaturated P2 building scaffold wherein Rq is other than hydrogen,such as a methylated cyclopentene scaffold can be prepared as shown inscheme 11.

Oxidation of commercially available 3-methyl-3-buten-1-ol (11a) by theuse of an oxidation agent like pyridinium chlorochromate followed bytreatment with acetyl chloride, bromine and methanol provides theα-bromo eater (11c). The afforded ester (11c) can then be reacted withthe enolate (11e), achieved for example by treatment of thecorresponding tert-butyl ester with a base such as lithium diisopropylamide in a solvent like tetrahydrofuran, to give the alkylated compound(11f). The tert-butyl ester (11e) can be prepared by treatment of thecorresponding commercially available acid (11d) where k′ is 1 to 3 withdi-tert-butyl dicarbonate in the presence of a base likedimethylaminopyridine. Cyclisation of (11f) by an olefin metathesisreaction performed as described above provides cyclopentene derivative(11g). Stereoselective epoxidation of (11g) can be carried out using theJacobsen asymmetric epoxidation method to furnish the epoxide (11 h).Finally, addition of a base like DBN (1,5-diazabicyclo-[4.3.0]non-5-ene)yields the alcohol (11i). Optionally the double bond of compound (11i)can be reduced for example by catalytic hydrogenation using a catalystlike palladium on carbon which provides the corresponding saturatedcompound.

The afforded cyclic scaffolds can then be used, as described above, tocomplete the inhibitor sequence. An example is shown in scheme 12.

The amino group of a P1-building block or a suitable precursor thereof(12b) can be coupled to the acid of the cyclopentene derivative (12a)using standard amide coupling conditions such as using HATU in thepresence of a base like diisopropyl phenylamine or the like, followed byintroduction of the R⁸-substituent for example by Mitsunobu conditionsas described above to provide (12d). Hydrolysis of the remaining esterand subsequent amide coupling of a desired P3 or P3-P4 building block(12e) optionally followed by manipulations of the P1 part providescyclopentene containing compounds (12f) according to general formula VI.When R⁷, R^(7′) and A′ contain functional groups, these are optionallysuitably protected by methods recognized by persons sidled in the art,see for example Bodanzky or Greene cited above.

Compounds having a hydrazine containing capping group attached directlyto the P2 moiety, i.e. P3 and P4 are absent and G is NRjNRj, can beprepared as depicted in Scheme 13

Reaction of tert-butyl carbazate (13a), optionally alkyl substituted onone or both nitrogens, with the add (13b) under peptide couplingconditions like with HATU and DIEA in a solvent like DMF provides 9Ac.Optional removal of the boc group by standard procedures like acidictreatment with for example TFA in a suitable solvent such asdichloromethane, provides the hydrazine containing derivative (13d).Alternatively, any appropriate hydrazine derivative, such asmorpholin-1-ylamine, piperidin-1-ylamine or the like can be linked tothe acid (13b) instead of the tert-butyl carbazate derivative.

The achieved compound can then be further extended by coupling of a P3or P4-P3 building block to the primary amine of compound 13d for examplems shown in scheme 14.

Treatment of the α-amino compound (14a) with sodium nitrite, potassiumbromide and sulphuric acid (Yang et al. J. Org. Chem. (2001), 66,7303-7312) provides the corresponding α-bromo compound (14b) which uponreaction with the above described derivative (13d) provides thehydrazine containing derivative (14c).

Compounds lacking a carboxy group in the P3 unit can be prepared asillustrated in Scheme 15 exemplified with a cyclopentane derivative asP2 scaffold.

The acid (15a) can be coupled to an amino azide derivative (15b),prepared by methods known from the literature using standard peptidecoupling conditions to give the amide derivative (15c). Reduction of theazide function for example by polymer bound triphenyl phosphine in asolvent like methanol or any other suitable reduction method providesintermediate (15d) which subsequently can be reacted with an acid underpeptide coupling conditions or with an amine in a reductive aminationreaction providing amides and secondary amines respectively.

Scheme 16 shows an alternative route towards compounds lacking a carboxygroup in the P3 unit

Instead of using the azide derivative (15b) in scheme 15 thecorresponding, optionally protected, hydroxy derivative (16b) can beused in the coupling with the acid (16a) and thus introducing a primaryalcohol. The alcohol (16c) can then, after optional deprotection, beoxidized with a suitable oxidizing agent like for example Dess-Martinperiodinane to form the corresponding aldehyde. Reaction of the aldehydewith a desired amine in a reductive amination reaction using a reagentlike for example polystyrene bound cyanoborohydride in a solvent likeTHF provides amine derivatives (16e).

Alternatively alcohol (16c) can be reacted with a suitable acylating oralkylating agent under the appropriate conditions to provide ester andether compounds respectively, i.e. G is O in general formula VI.

Subsequent reaction of the formed alcohol with a suitable acylating oralkylating agent using the appropriate conditions provides the ester andether compounds respectively, i.e. G is O in general formula VI.

Although Scheme 15 and 16 have been described with reference to acyclopentane derivative i.e. q′ is 0 and k is 1 in Formula VI, it willbe readily apparent that the corresponding methodology is applicable forother compounds of the Formula VI.

When R⁷, R^(7′) and A′ contains functional groups, these are suitablyprotected by methods recognized by persons skilled in the art, see forexample Bodanzky or Greene cited above.

Formation of Macrocyclic Compounds

Compounds according to the present invention wherein an alkylene chainextending from the R⁷/R^(7′) cycloalkyl to Rx, Rd or R¹¹ thus forming amacrocycle, can be prepared as described below. Suitable P1, P2 and P3building blocks, or precursors thereof, are coupled together using thestrategies described above, followed by a ring-closing reaction(macrocyclization). The substituent W—R⁸ of the P2 building block can beincorporated via a Mitsunobu reaction as described above, before orafter formation of the macrocycle or the assembly can be done with therequired substituted proline analogue or carbocycle. For macrocyclicstructures extending from the R⁷/R^(7′) cycloakyl to R¹¹, P3 amino acidscontaining the appropriate side chain can be prepared as described inWO00/59929.

A typical route to macrocyclic compounds is shown in Scheme 17 whichillustrates the technique applied to a compound having aspiro-cyclopropyl P1, where the macrocycle incorporates the P3 sidechain.

Coupling of acid derivative (17a) with the appropriate, acid protected,amino acid (17b) using standard peptide coupling conditions as describedabove provides (17c). Formation of the macrocycle can then be carriedout via an olefin metathesis reaction using a Ru-based catalyst such asthe one reported by Miler, S. J., Blackwell, H. E.; Grubbs, R. H. J. Am.Chem. Soc. 118, (1996), 9606-9614, Kingsbury, J. S., Harrity, J. P. A.,Bonitatebus, P. J., Hoveyda, A. H., J. Am. Chem. Soc. 121, (1999),791-799 and Huang et al., J. Am. Chem. Soc. 121, (1999), 2674-2678. Itwill also be recognized that catalysts containing other transitionmetals such as Mo can be used for this reaction. Optionally the doublebond is reduced and/or the ethyl ester is hydrolysed by standardhydrogenation and/or hydrolysation methods respectively well known inthe art. Alternatively the methyl ester can be selectively hydrolysedfollowed by coupling of a R¹⁸-G-P4 building block by standard peptidecoupling conditions. The macrocyclisation step described in Scheme 17can also be applied to the corresponding carbocyclic analogues describedabove. When the linker contains a nitrogen atom the ring closure can becarried out by reductive amination as described in WO00/59929.

Macrocyclic compounds without the cyclopropyl moiety in the P1 part, i.ethe macrocyclic ring extends directly from the peptidic backbone at thecarbon adjacent R⁷, can be prepared using the method described herein.An example wherein a 5 membered cycloakyl derivative is used as the P2scaffold is shown in scheme 18.

Coupling of a suitable allylglycine derivative (18a), to the acidfunction of the P2 scaffold (18b) using standard peptide couplingconditions yields the amide derivative (18c). Hydrolysis of the estergroup followed by a peptide coupling reaction with the olefinsubstituted amino acid (18Ad) provides the amide compound (18e). A ringclosing metathesis reaction is then effected by using for exampleHoveyda-Grubbs catalyst which gives the macrocyclic compound (18f).

Even though scheme 18 shows the synthetic sequence using a P2 scaffoldwith an unsubstituted hydroxy group, it will be apparent that the R8substituent can be introduced at any convenient stage of the synthesisfor example as described in scheme 9 and 10 or it can be introducedafter the metathesis reaction, i.e. on compound 18f, using any of themethods described herein.

Building blocks to be used in the preparation of compounds wherein themacrocycle extends from the amide nitrogen in the P3 fragment i.e Rx isJ in general formula VI, or in the preparation of compounds wherein theP3 and P4 fragments are absent, i.e. m and n are 0 and G is NRj ingeneral formula VI, can typically be prepared as outlined in scheme 18B.

Carbamate 18Ba, which is commercially available or is readily preparedfor instance by reaction of the desired alkyl amine with di-tert-butyldicarbonate, can be reacted with an appropriate ω-unsaturated alcoholunder Mitsunobu conditions to provide the alkylated carbamate (18Bb).Subjection of 188b to acidic conditions like for example treatment withtrifluoroacetic acid in a solvent like dichloromethane gives the freeamine (18Bc) which can be linked to a P2 fragment using any of thepreviously desorbed strategies.

Macrocyclic structures containing a hydrazine group i.e. T is NRd or mand n are 0 and G is NRjNRj, in general formula VI, can be prepared bylinking a suitably N-alkylated carbazate derivative to the P2 fragment.Alkylated carbazate derivatives can be prepared, for example, asdescribed in Scheme 19.

Oxidation of the appropriate alcohol (19a) effected by a suitableoxidation method like for example with N-methyl morpholine oxide andtetrapropylammonium perruthenate in a solvent like dichloromethaneprovides aldehyde (19b). Reductive alkylation of tert-butyl carbazatewith the afforded aldehyde gives the desired N-alkylated building block(19c). Alternatively, any desired hydrazine derivative such asmorpholin-1-ylamine, piperidin-1-ylamine or the like can be used insteadof tert-butyl carbazate in the reaction with aldehyde 19b.

Scheme 20 illustrates synthetic sequences to building blocks suitablefor the preparation of compounds wherein the “outer” nitrogen of thehydrazine group is alkylated, either with an w-unsaturated alkyl chainappropriate for subsequent macrocycle formation or with any othersuitable alkyl group.

Reaction of a suitably protected hydrazine derivative, for example(1,3-dioxo-1,3-dihydro-isoindol-2-yl)-carbamic acid tert-butyl ester(20a), which can easily be prepared by a person skilled in the art, witha desired alcohol, R—OH, under Mitsunobu conditions provides N-alkylatedhydrazine compound (20b). Removal of the phtalimido group effected bytreatment with hydrazine or a derivative thereof like hydrazine hydrateor hydrazine acetate provides the carbazate (20c). The afforded primaryamine can then either be be coupled to any desired P2 fragment using anyof the methods previously described to give 20d or alternatively it canbe further alkylated using for example the reductive amination methoddescribed in scheme 19 followed by coupling to a P2 fragment aspreviously described to give 20.

Scheme 21 exemplifies the coupling of a hydrazine containing P3 buildingblock to a cyclopentane scaffold followed by macrocyclisation.

Coupling of the carbazate derivative (21b) to the P2-P1 building block(21a) using standard peptide coupling conditions provides intermediate(21c). Ring closure of (21c) by an olefin metathesis reaction asdescribed in scheme 18 gives the macrocyclic compound (21d).

The term “N-protecting group” or “N-protected” as used herein refers tothose groups intended to protect the N-terminus of an amino acid orpeptide or to protect an amino group against undesirable reactionsduring synthetic procedures. Commonly used N-protecting groups aredisclosed in Greene, “Protective Groups in Organic Synthesis” (JohnWiley & Sons, New York, 1981), which is hereby incorporated byreference. N-protecting groups include acyl groups such as formyl,acetyl, propionyl, pivaloyl, t-butylacetyl, 2-chloroacetyl,2-bromoacetyl, trifluoracetyl, trichloroacetyl, phthalyl,o-nitrophenoxyacetyl, α-chlorobutyryl, benzoyl, 4-chlorobenzoyl,4bromobenzoyl, 4-nitrobenzoyl, and the like; sulfonyl groups such asbenzenesulfonyl, p-toluenesulfonyl, and the like, carbamate forminggroups such as benzyloxycarbonyl, p-chlorobenzyl-oxycarbonyl,p-methoxybenzyloxycarbonyl, p-nitrobenzyloxycarbonyl,2-nitrobenzyloxycarbonyl, p-bromobenzyloxycarbonyl,3,4-dimethoxybenzyloxycarbonyl, 4-methoxybenzyloxycarbonyl,2-nitro-4,5-dimethoxybenzyloxycarbonyl,3,4,5-trimethoxybenzyloxycarbonyl,1-(p-biphenylyl)-1-methylethoxycarbonyl,α,α-dimethyl-3,5-dimethoxybenzyloxycarbonyl, benzhydryloxycarbonyl,t-butoxycarbonyl, diisopropylmethoxycarbonyl, isopropyloxycarbonyl,ethoxycarbonyl, methoxycarbonyl, allyloxycarbonyl,2,2,2-trichloroethoxycarbonyl, phenoxycarbonyl, 4-nitrophenoxycarbonyl,fluorenyl-9-methoxycarbonyl, cyclopentyloxycarbonyl,adamantyloxycarbonyl, cyclohexyloxycarbonyl, phenylthiocarbonyl, and thelike; alkyl groups such as benzyl, triphenylmethyl, benzyloxymethyl andthe like; and silyl groups such as trimethylsilyl and the like. FavouredN-protecting groups include Fmoc, formyl, acetyl, benzoyl, pivaloyl,t-butylacetyl, phenylsulfonyl, benzyl, t-butoxycarbonyl (BOC) andbenzyloxycarbonyl (Cbz).

Hydroxy protecting group as used herein refers to a substituent whichprotects hydroxyl groups against undesirable reactions during syntheticprocedures such as those O-protecting groups disclosed in Greene,“Protective Groups in Organic Synthesis,” (John Wiley & Sons, New York(1981)). Hydroxy protecting groups comprise substituted methyl ethers,for example, methoxymethyl, benzyloxymethyl, 2-methoxyethoxymethyl,2-(trimethylsilyl)ethoxymethyl, t-butyl and other lower alkyl ethers,such as isopropyl, ethyl and especially methyl, benzyl andtriphenylmethyl; tetrahydropyranyl ethers; substituted ethyl ethers, forexample, 2,2,2-trichloroethyl; silyl ethers, for example,trimethylsilyl, t-butyldimethylsilyl and t-butyldiphenylsilyl; andesters prepared by reacting the hydroxyl group with a carboxylic acid,for example, acetate, propionate, benzoate and the like.

In treating conditions caused by flavivirus such as HCV, the compoundsof formula VI are typically administered in an amount to achieve aplasma level of around 100 to 5000 nM, such as 300 to 2000 nM. Thiscorresponds to a dosage rate, depending on the bioavailability of theformulation, of the order 0.01 to 10 mg/kg/day, preferably 0.1 to 2mg/kg/day. A typical dosage rate for a normal adult will be around 0.05to 5 g per day, preferably 0.1 to 2 g such as 500-750 mg, in one to fourdosage units per day. As with al pharmaceuticals, dosage rates will varywith the size and metabolic condition of the patient as well as theseverity of the infection and may need to be adjusted for concomitantmedications.

As is good prescribing practice with antiviral therapy, the compounds offormula I are typically coadministered with other HCV therapies to avoidthe generation of drug escape mutants. Examples of such additional HCVantiviral therapies include ribavirin, interferons, including pegylatedinterferons. Additionally a number of nucleoside analogues and proteaseinhibitors are in clinical or preclinical development and will beamenable to co-administration with the compounds of the invention.

Accordingly a further aspect of the invention provides a compositioncomprising a compound of formula I and at least one further HCVantiviral in a common dosage unit, such as any of the dosage formsdescribed below, but especially an orally administered tablet, orcapsule or a liquid suspension or solution for oral or injection use. Afurther aspect of the invention provides a method for the treatment orprophylaxis of flavivirus infection, such as HCV, comprising thesequential or simultaneous administration of a compound of formula I andat least one further HCV antiviral. A related aspect of the inventionprovides a patient pack comprising a first pharmaceutical composition,preferably in unit dosage form, of the compound of formula I and asecond pharmaceutical composition of a second HCV antiviral, typicallyalso in unit dosage form and generally in a separate container withinthe patient pack. A patient pack will conveniently also be provided withinstructions printed on the package or a container therein, or on apackage insert, for the simultaneous or sequential administration of therespective pharmaceutical compositions.

Many HCV patients are co-Infected, or prone to superinfection, withother infectious diseases. Accordingly, a further aspect of theinvention provides combination therapies comprising the compound of theinvention co-formulated in the same dosage unit or co-packaged with atleast one further anti-infective pharmaceutical. The compound of theinvention and the at least one further antinfective are administeredsimultaneously or sequentially, typically at doses corresponding to themonotherapy dose for the agent concerned. However, certain antifectivescan induce a synergistic response, allowing one or both of the activeingredients to be administered at a lower dose that the correspondingmonotherapy. For example in drugs prone to rapid metabolism by Cyp3A4,co-dosing with the HIV protease inhibitor ritonavir can allow lowerdosage regimes to be administered.

Typical coinfections or superinfections with HCV include hepatitis Bvirus or HIV. Accordingly the compound of the invention isadvantageously co-administered (either in the same dosage unit,co-packaged or separately prescribed dosage unit) with at least one HIVantiviral and/or at least one HBV antiviral.

Representative HIV antivirals include NRTI such as alovudine (FLT),zudovudine (AZT, ZDV), stavudine (d4T, Zerit), zalcitabine (ddC),didanosine (ddl, Videx), abacavir, (ABC, Ziagen), lamivudine (3TC,Epivir), emtricitabine (FTC, Emtriva), racevir (racemic FTC), adefovir(ADV), entacavir (BMS 200475), alovudine (FLT), tenofovir disoproxilfumarate (TNF, Viread), amdoxavir (DAPD), D-d4FC (DPC-817), -dOTC (ShireSPD754), elvucitabine (Achllion ACH-126443), BCH 10681 (Shire) SPD-756,racivir, D-FDOC, G07340, iNK-20 (thioether phospholipid AZT, Kucera),2′3′-dideoxy-3′-fluoroguanosine (FLG) & its prodrugs such as MIV-210,reverset (RVT, D-D4FC, Pharmasset DPC-817).

Representative NNRTI include delavirdine (Rescriptor), efavirenz(DMP-266, Sustiva), nevirapine (BIRG-587, Viramune), (+)calanolide A andB (Advanced Life Sciences), capravirine (AG1549f S-1153; Pfizer),GW-695634 (GW-8248; GSK), MIV-150 (Medivir), MV026048 (R-1495; MedivirAB/Roche), NV-05 2 2 (Idenix Pharm.), R-278474 (Johnson & Johnson),RS-1588 (Idenix Pharm.), TMC-120/125 (Johnson & Johnson), TMC-125(R-165335; Johnson & Johnson), UC-781 (Biosyn Inc.) and YM215389(Yamanoushi).

Representative HIV protease inhibitors include PA-457 (Panacos), KPC-2(Kucera Pharm.), 5 HGTV-43 (Enzo Biochem), amprenavir (VX-478,Agenerase), atazanavir (Reyataz), indinavir sulfate (MK-639, Crixvan),Lexiva (fosamprenavir calcium, GW-433908 or 908, VX-175), ritonavir(Norvir), lopinavir+ritonavir (ABT-378, Kaletra), tipranavir, nelfinavirmesylate (Viracept), saquinavir (Invirase, Fortovase), AG1776 (JE-2147,KNI-784; Nippon Mining Holdings), AG-1859 (Pfizer), DPC-681/684 (BMS),GS224338; Gilead Sciences), KNI-272 (Nippon Mining Holdings), Nar-DG-35(Narhex), P(PL)-100 (P-1946; Procyon Biopharma), P-1946 (ProcyonBiopharma), R-944 (Hoffmann-LaRoche), RO-0334649 (Hoffmann-LaRoche),TMC-114 (Johnson & Johnson), VX-385 (GW40385; GSK/Vertex), VX-478(Vertex/GSK).

Other HIV antivirals include entry inhibitors, including fusioninhibitors, inhibitors of the CD4 receptor, inhibitors of the CCR5co-receptor and inhibitors of the CXCR4 coreceptor, or apharmaceutically acceptable salt or prodrug thereof. Examples of entryinhibitors are AMD-070 (AMD11070; AnorMed), BlockAlde/CR (ADVENTRXPharm.), BMS 806 (BMS-378806; BMS), Enfurvirtide (T-20, R698, Fuzeon),KRH1636 (Kureha Pharmaceuticals), ONO-4128 (GW-873140, AK-602, E-913;ONO Pharmaceuticals), Pro-140 (Progenics Pharm), PRO542 (ProgenicsPharm.), SCH-D (SCH-417690; Schering-Plough), T-1249 (R724;Roche/Trimeris), TAK-220 (Takeda Chem. Ind.), TNX-355 (Tanox) andUK-427,857 (Pfizer). Examples of integrase inhibitors are L-870810(Merck & Co.), c-2507 (Merck & Co.) and S(RSC)-1838 (shionogi/GSK).

Examples of HBV antivirals include adefovir dipivoxil (Hepsera), andespecially lamivudine and 2′3′-dideoxy-3′-fluoroguanosine (FLG) & itsprodrugs such as MIV-210, the 5′-O-valyl-L-lactyl prodrug of FLG. Theselatter HBV antivirals are particularly convenient as they are alsoactive against HIV.

While it is possible for the active agent to be administered alone, itis preferable to present it as part of a pharmaceutical formulation.Such a formulation will comprise the above defined active agent togetherwith one or more acceptable carriers or excipients and optionally othertherapeutic ingredients. The carrier(s) must be acceptable in the senseof being compatible with the other ingredients of the formulation andnot deleterious to the recipient.

The formulations include those suitable for rectal, nasal, topical(including buccal and sublingual), vaginal or parenteral (Includingsubcutaneous, intramuscular, intravenous and intradermal)administration, but preferably the formulation is an orally administeredformulation. The formulations may conveniently be presented in unitdosage form, e.g. tablets and sustained release capsules, and may beprepared by any methods well known in the art of pharmacy.

Such methods include the step of bringing into association the abovedefined active agent with the carrier. In general, the formulations areprepared by uniformly and Intimately bringing into association theactive agent with liquid carriers or finely divided sold carriers orboth, and then if necessary shaping the product. The invention extendsto methods for preparing a pharmaceutical composition comprisingbringing a compound of Formula VI or its pharmaceutically acceptablesalt in conjunction or association with a pharmaceutically acceptablecarrier or vehicle. If the manufacture of pharmaceutical formulationsinvolves intimate mixing of pharmaceutical excipients and the activeingredient in salt form, then it is often preferred to use excipientswhich are non-basic in nature, i.e. either acidic or neutral.Formulations for oral administration in the present invention may bepresented as discrete units such as capsules, cachets or tablets eachcontaining a predetermined amount of the active agent; as a powder orgranules; as a solution or a suspension of the active agent in anaqueous liquid or a non-aqueous liquid; or as an oil-n-water liquidemulsion or a water in oil liquid emulsion and as a bolus etc.

With regard to compositions for oral administration (e.g. tablets andcapsules), the term suitable carrier includes vehicles such as commonexcipients e.g. binding agents, for example syrup, acacia, gelatin,sorbitol, tragacanth, polyvinylpyrrolidone (Povidone), methylcellulose,ethylcellulose, sodium carboxymethylcellulose,hydroxypropylmethylcellulose, sucrose and starch; fillers and carriers,for example corn starch, gelatin, lactose, sucrose, microcrystallinecellulose, kaolin, mannitol, dicalcium phosphate, sodium chloride andalginic add; and lubricants such as magnesium stearate, sodium stearateand other metallic stearates, stearic acid, glycerol stearate, siliconefluid, talc waxes, oils and colloidal silica. Flavouring agents such aspeppermint, oil of wintergreen, cherry flavouring or the like can alsobe used. It may be desirable to add a colouring agent to make the dosageform readily identifiable. Tablets may also be coated by methods wellknown in the art.

A tablet may be made by compression or moulding, optionally with one ormore accessory ingredients. Compressed tablets may be prepared bycompressing in a suitable machine the active agent in a free flowingform such as a powder or granules, optionally mixed with a binder,lubricant, inert diluent, preservative, surface-active or dispersingagent. Moulded tablets may be made by moulding in a suitable machine amixture of the powdered compound moistened with an inert liquid diluent.The tablets may be optionally be coated or scored and may be formulatedso as to provide slow or controlled release of the active agent.

Other formulations suitable for oral administration include lozengescomprising the active agent in a flavoured base, usually sucrose andacacia or tragacanth; pastilles comprising the active agent in an inertbase such as gelatin and glycerin, or sucrose and acacia; andmouthwashes comprising the active agent in a suitable liquid carrier.

The compounds of formula VI can form salts which form an additionalaspect of the invention. Appropriate pharmaceutically acceptable saltsof the compounds of formula I include salts of organic acids, especiallycarboxylic acids, including but not limited to acetate,trifluoroacetate, lactate, gluconate, citrate, tartrate, maleate,malate, pantothenate, isethionate, adipate, alginate, aspartate,benzoate, butyrate, digluconate, cyclopentanate, glucoheptanate,glycerophosphate, oxalate, heptanoate, hexanoate, fumarate, nicotinate,palmoate, pectinate, 3-phenylpropionate, picrate, pivalate, proprionate,tartrate, lactobionate, pivolate, camphorate, undecanoate and succinate,organic sulphonic acids such as methanesulphonate, ethanesulphonate,2-hydroxyethane sulphonate, camphorsulphonate, 2-napthalenesulphonate,benzenesulphonate, p-chlorobenzenesulphonate and p-toluenesulphonate;and inorganic acids such as hydrochloride, hydrobromide, hydroiodide,sulphate, bisulphate, hemisulphate, thiocyanate, persulphate, phosphoricand sulphonic acids. The invention further extends to salts of thecompounds of formula I which may or may not be pharmaceuticallyacceptable, but which are useful as synthetic intermediates, the saltmoiety being displaced or replaced as necessary.

The invention includes prodrugs of the compounds of formula I. Prodrugsof the compounds of formula VI are those compounds which followingadministration to a patient release a compound of the formula VI in vivogenerally following hydrolysis in the gut, liver or plasma. Typicalprodrugs are pharmaceutically acceptable ethers and especially esters(including phosphate esters) of hydroxy functions, pharmaceuticallyacceptable amides or carbamates of amine functions or pharmaceuticallyacceptable esters of carboxy functions. Preferred pharmaceuticallyacceptable esters include alkyl esters, including acetyl, ethanoyl,butyryl, t-butyryl, stearyl and pivaloyl, phosphate esters and sulphonicesters (ie those derived from RSO₂OH, where R is lower alkyl or aryl).Pharmaceutically acceptable esters include lower alkyl ethers and theethers disclosed in WO00/47561, especially methoxyaminoacyl andethoxyaminoacyl.

The compounds of the invention have various steric centres and theinvention extends to racemates and enantiomers at each of these stericcentres.

Typically, the stereochemistry of the groups corresponding to the P3 andP4 side chains (ie R¹⁵ and/or R¹¹) will correspond to an L-amino acidconfiguration, although the invention also extends to D-isomers at oneor both of these centres. It is noteworthy that the L configuration isactive notwithstanding that the nature of the E moiety means that P3 andP4 are typically translated one atom relative to a conventionalpolypeptide and the fact that reversal of a peptide residue, asenvisaged for P3 and P4 then pitches the amine acid side chain to theopposite side compared to a conventional peptide substrate.

The stereochemistry of the backbone component of the cyclic P2 group(i.e. spanning the carbonyl of the P1 amide bond and the carbonylextending of P3 will typically correspond to L-proline. Thestereochemistry of the P2 ring atom to which W is bonded is typically asshown:

In compounds of the invention wherein R⁷ and R^(7′) together define aspiroalkyl group, such a spiro-cycloakyl will typically comprise an Rsubstituent on the spiro-cyclopropyl ring which is is orientated syn toA

or anti to A:

Conveniently, the spiro carbon of such a spiro-cyclopropyl ring has theR configuration:

Conveniently an R^(7′a) substituent on a spiro-cyclopropyl ring adjacentto A is in a syn orientation in the following absolute configuration:

Particularly preferred variants have R^(7′a) include ethyl, hence theasymmetric carbon atoms at position 1 and 2 have the R. R configuration.Alternative preferred R Include vinyl, hence the asymmetric carbon atomsat position 1 and 2 have the R, S configuration.

Where the compound of the invention is a macrocycle comprising a Jgroup, J is preferably a diastereomer represented by partial structures(i) or (ii):

especially where J is syn to A.

DETAILED DESCRIPTION OF THE EMBODIMENTS

Various embodiments of the invention will now be described by way ofillustration only with reference to the following non-limiting examples.

Example 1

7-Methoxy-2-phenyl-quinolin-4-ol (1)

To a stirred round bottled flask with toluene (100 mL) ethyl benzoylacetate (18.7 g, 97 mmol) and m-anisidine (12 g, 97 mmol) was added. 4 MHCl in dioxane (0.5 mL) was added and the reaction mixture was refluxedfor 6 h (140° C.). The mixture was co-evaporated with toluene. To thecrude mixture diphenyl ether (50 mL) was added and the mixture washeated to 280° C. for 2 h. When the theoretical amount ethanol (6 mL)was collected in a Dean Stark trap the heating was stopped and themixture was cooled to rt. The crude mixture was dissolved in CH₂Cl₂ (100mL) and stirred for 30 min. The formed precipitate was filtered off anddried which gave 1 (4.12 g, 16.4 mmol, 17%): pale yellow powder.

¹H (300 MHz, DMSO-D₆): δ 3.8 (s, 3H), 6.24 (s, 1H), 6.88-6.96 (dd, 1H,J=9.07 Hz, J=2.47 Hz), 7.19 (d, 1H, J=2.19 Hz), 7.56 (t, 3H, J=2.19 Hz),7.8 (dd, 2H, J=7.14 Hz, J=2.19 Hz), 8.0 (d, 1H, J=9.06 Hz); ¹³C (75.5MHz, DMSO-D₆): δ 55.3, 99.6, 106.9, 113.1, 119.1, 126.4, 127.5, 128.8,130.2, 134.1, 142.2, 149.4, 161.8, 176.4.

Example 2

(Rac)-4-oxocyclopent-2-ene-1, 2-dicarboxylic acid dimethyl ester (2)

(1R,2S)-4-oxo-cyclopentane-1, 2-dicarboxylic acid dimethyl ester (4.8 g,23.8 mmol) and CuBr₂ (11.9 g, 532 mmol) were dissolved in dry THF (70mL) and the mixture was refluxed for two hours at 90° C. The formed CuBrwas filtrated off and the organic phase was concentrated. CaCO₃ (2.7 g,27.2 mmol) and DMF (70 mL) were added and the mixture was held at 100°C. for one hour. The dark brown mixture was poured over ice (35 g) andthe formed precipitate was filtrated off. The aqueous layer wasextracted with ethyl acetate (1×300 mL+3×150 mL). The organic phaseswere dried, filtrated and concentrated. Purification by flashchromatography (toluene/EtOAc 9:1) gave 2 (2.1 g, 45%) as yellowcrystals

Example 3

((1S,4R) & (1R,4S))-4-hydroxy-cyclopent-2-ene-1,2-dicarboxylic adddimethyl ester (3)

To a cold solution (−30° C.) of 2 (3.18 g, 16.1 mmol) dissolved in MeOH(23 mL), NaBH₄ (0.66 g, 17.5 mmol) was added. After nine minutes theexcess of NaBH₄ was destroyed by adding brine (80 mL). The mixture wasconcentrated and extracted with ethyl acetate (4×80 mL). The organicphases were dried, filtrated and concentrated and gave 3 (3.0 g, 92%) asa yellow oil.

Example 4

(1 S,4R) & (1R,4S)-4-hydroxy-cyclopent-2-ene-1,2-dicarboxylic add2-methyl ester (4)

To an ice-cold solution of 3 (3.4 g, 22 mmol) dissolved in dioxane andwater (1:1, 110 mL), LiOH (0.52 g, 22 mmol) was added. After two and ahalf hours the mixture was co-evaporated with toluene and methanol.Purification by flash chromatography (toluene/Ethyl acetate 3:1+1% HOAc)gave the title compound (1.0 g, 27%) as yellow-white crystals.

¹H-NMR (300 MHz, CD₃OD): δ 1.78-1.89 (m, 1H), 2.70-2.84 (m, 1H),3.56-3.71 (m, 1H), 3.76 (s, 3H), 4.81-4.90 (m, 1H), 6.76-6.81 (m, 1H);¹³C-NMR (75.5 MHz, CDCl₃): δ 38.0, 48.0, 52.4, 75.7, 137.0, 148.2, 165.0178.4.

Example 5

((3S,5R) &(3R,5S))-5-((S)-1-tert-Butoxycarbonyl-butylcarbamoyl)-3-hydroxy-cyclopent-1-enecarboxylicacid methyl (5)

To an ice cooled solution of 4 (0.20 g, 1.1 mmol) and 2-amino-pentanoicadd tert.butyl ester (0.24 g, 1.4 mmol) in DMF (7 mL), DIPEA (0.18 g,1.4 mmol) and HATU (0.53 g, 1.4 mmol) were added. After two hours thesolution was concentrated and purified using column chromatography(toluene/ethyl acetate 3:1). This gave the title compound as a yellowoil (0.22 g, 63%).

¹H-NMR (300 MHz, CDCl₃): δ 0.84-0.96 (m, 3H), 1.14-1.39 (m, 2H), [(1.44& 1.49) s, 9H], 1.50-1.60 (m, 1H), 1.61-1.85 (m, 1H), 1.97-2.10 (m, 1H),2.11-2.28 (m, 1H), 3.57-3.68 (m, 1H), [(3.73 & 3.76) s, 3H], 4.30-4.50(m, 1H), 4.63-4.73 (m, 1H), 6.8068.95 (m, 1H), 6.95-7.00 (m, 1H).

Example 6

((3S,5R) &(3R,5S))-5-((S)-1-tert-Butoxycarbonyl-propylcarbamoyl-3-hydroxy-cyclopent-1-necarboxylicacid methyl ester (6)

Reaction of 4 (141 mg, 76 mmol) according to the method described forthe preparation of 5 using L-2-amino-N-butyric acid tert.butyl esterinstead of 2-amino-pentanoic acid tert.butyl ester gave the titlecompound as a slightly yellow oil (171 mg, 69%).

¹H-NMR (300 MHz, CDCl₃): δ 0.89-0.98 (m, 3H), [(1.42 & 1.44) s, H],1.60-1.78 (m, 1H), 1.79-1.95 (m, 1H), 1.99-2.11 (m, 1H), 2.18-2.30 (m,1H), 3.58-3.65 (m, 1H), [3.75 & 3.78) s, 3H], 4.22-4.39 (m, 1H),4.61-4.66 (m, 1H), 6.77-6.90 (m, 1H), 6.91-6.92 (m, 1H).

Example 7

((3S,5R) &(3R,6S′))-5-((1R,2S)-1-tert-Butoxycarbonyl-2-vinyl-cyclopropylcarbamoyl)-3-hydroxy-cyclopent-1-enecarboxylicacid methyl ester (7)

Reaction of 4 (50 mg, 37 mmol) according to the method described for thepreparation of 5 using (1R, 2S)-1-amino-2-vinyl-cyclopropane carboxylicacid tert.butyl ester instead of 2-amino-pentanoi acid tert.butyl esterprovided the title compound as a slightly yellow oil (650 mg, 38%).

¹H-NMR (300 MHz, CDCl₃): δ [(1.38 & 1.42) s, 9H], 1.75-1.83 (m, 1H),2.00-2.21 (m, 3H), 3.55-3.63 (m, 1H), [(3.77 & 3.82) s, 3H], 4.20-4.38(m, 1H), 4.65-4.80 (m, 1H), 5.13-5.20 (m, 1H), 5.22-5.38 (m, 1H),5.60-6.82 (m, 1H), 6.95-8.96 (m, 2H).

Example 8

((3R,5R) &(3S,5S))-5-((S)-1-tert-Butoxycarbonyl-butylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarboxylicacid methyl ester (8)

To an ice cooled solution of 5 (0.23 g, 0.67 mmol) in dry THF,7-methoxy-2-phenyl-quinolin-4-ol (0.22 g, 0.88 mmol) andtriphenylphosphine (0.23 g, 0.88 mmol) were added. Then DIAD (0.19 g,0.92 mmol) was dissolved in THF (2 mL) and added dropwise to thesolution. After one hour the mixture was concentrated and purified usingflash chromatography (toluene/ethyl acetate 3:1). This gave the titlecompound as a white powder (0.30 g, 77%).

¹H-NMR (300 MHz, CDCl₃): δ 0.88-1.00 (m, 3H), 1.18-1.43 (m, 2H), [(1.45& 1.50) s, 9H], 1.53-1.65 (m, 1H), 1.68-1.85 (m, 1H), 2.29-2.43 (m, 1H),3.10-3.25 (m, 1H), [(3.79 & 3.83) s, 3H], 3.97 (s, 3H), 4.05-4.20 (m,1H), 4.38-4.50 (m, 1H), 6.03-6.13 (m, 1H), 6.65-8.90 (m, 1H), 7.04-7.18(m, 3H), 7.40-7.56 (m, 4H), 8.00-8.12 (m, 3H).

Example 9

((3R,5R) &(3S,5S))-5-((S)-1-tert-Butoxycarbonyl-propylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarboxylicacid methyl ester (9)

Reaction of 6 (132 mg, 40 mmol) according to the method described forthe preparation of 8 gave the title compound as a yellow oil (137 mg,61%).

¹H-NMR (300 MHz, CDCl₃): δ 0.83-0.98 (m, 3H), [(1.42 & 1.44) a, 9H],1.65-1.78 (m, 1H), 1.80-1.97 (m, 1H), 2.30-2.40 (m, 1H), 3.05-3.20 (m,1H), [(3.78 & 3.80) s, 3H], 3.94 (s, 3H), 3.95-4.01 (m, 1H), 4.38-4.44(s, 1H), 6.05-6.15 (m, 1H), 6.80-6.94 (m, 1H), 7.02-7.15 (m, 3H),7.38-7.55 (m, 4H), 7.97-8.18 (m, 3H).

Example 10

((3R,5R) &(3S,5S))-5-((1R,2S)-1-tert-Butoxycarbonyl-2-vinyl-cyclopropylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarboxylicacid methyl ester (10)

Reaction of 7 (41 mg, 116 mmol) according to the method described forthe preparation of 8 provided the title compound as a yellow oil.

¹H-NMR (300 MHz, CDCl₃): δ 1.52-1.57 (m, 1H), 1.58 (m, 9H), 1.80-1.83(m, 1H), 2.00-2.17 (m, 1H), 2.20-2.38 (m, 1H), 3.20-3.37 (m, 1H), 3.80(s, 3H), 3.81-3-3.98 (m, 1H), 3.99 (s, 3H), 5.12-6.20 (m, 1H), 5.22-5.40(m, 1H), 5.63-5.80 (m, 1H), 6.05-6-20 (m, 1H), 7.00-7.21 (m, 4H),7.40-7.58 (m, 4H), 8.02-8.18 (m, 3H).

Example 11

((3R,5R) &(3S,5S)-5-((S-1-tert-Butoxycarbonyl-butylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarboxylicacid (11)

The methyl ester 8 (0.35 g, 0.61 mmol) was dissolved in dioxane/water(1:1, 7 mL) and LiOH (0.031 g, 1.3 mmol) was added. The reaction wasstirred over night and then co-concentrated. This gave the lithium saltof 11 (0.32 g, 90%) as a brown powder.

Example 12

((3R,5R)&(3S,5S))-5-((S)-1-tert-Butoxycarbonyl-propylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarboxylicacid (12)

Reaction of 9 (225 mg, 40 mmol) according to the method described forthe preparation of 11 provided the title compound as yellow salt (157mg, 72%).

Example 13

((3R,5R) &(3S,5S))-5-((1R,2S)-1-tert-Butoxycarbonyl-2-vinyl-cyclopropylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarboxylicAcid (13)

Reaction of 10 (35 mg, 59 mmol) according to the method described forthe preparation of 11 (33 mg, 97%) provided the title compound as ayellow salt.

Example 14

(S)-2-{[((1R,4S) &(1R,4R))-2-{(S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricacid tert-butyl ester (14)

The add 12 (38.4 mg, 0.070 mmol) and(2-amino-3-methyl-butyrylamino)-cyclohexyl acetic acid methyl ester(26.6 mg, 0.098 mmol) were dissolved in DMF (1.5 mL) and cooled in anice-bath. DIPEA (17.1 μL, 0.096 mmol) and HATU (37.4 mg, 0.098 mmol)were added. After ninety minutes the mixture was co-concentrated withtoluene and methanol and then purified by lash column chromatography(toluene/ethyl acetate 6:1). Further purification was performed on HPLC(90% MeOH+0.2% TEA). The diastereomeric mixture 14 was concentrated andgave a slightly yellow oil (20.6 mg, 37%). After lyophilisation 14 wascollected as a white powder. ¹H-NMR (300 MHz, CDCl₃): δ 0.93-1.02 (m,9H), 1.03-1.25 (m, 4H), 1.44 (s, 9H), 1.65-1.86 (m, 9H), 2.05-2.10 (m,1H), 2.22-2.40 (m, 1H), 3.05-3.20 (m, 1H), 3.77 (s, 3H), 3.98 (s, 3H),4.18-422 (m, 1H), 4.38-4.60 (m, 3H), 6.01-6.10 (m, 1H), 6.61-6.70 (m,2H), 6.80-6.85 (m, 1H), 7.05-7.18 (m, 2H), 7.40-7.58 (m, 5H), 8.00-8.13(m, 3H). ¹³C-NMR (75.5 MHz, CDCl₃): δ 9.7, 18.4, 19.2, (25.9 & 26.11,[28.2 & 28.5], 29.6, 32.0, 37.3, 41.0, 46.2, 50.7, 52.4, 54.4, 55.8,57.2, 58.5, 82.0, 82.8, 98.4, 110.2, 118.4, 120.1, 123.2, 127.9, 128.2,128.9, 129.5, 131.2, 135.1, 135.2, 142.7, 144.2, 161.6, 164.3, 164.7,170.9, 171.4, 172.4. MALDI-TOF m/z 821.56 [(M+Na)⁺ calcd forC₄₅H₅₈N₄NaO₉ ⁺ 821.41].

Example 15

(S)-2-[((1R,4R) & (1S,4S))-2-{(R)-1-[((R)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino)-butyricadd tert-butyl eater (15)

Reaction of 12 (20 mg, 37 mmol) according to the method described forthe preparation of 14 using(2-amino-3-methyl-butyrylamino)-(R)-cyclohexyl acetic acid methyl esterinstead of (2-amino-3-methyl-butyrylamino)-(S)-cyclohexyl acetic acidmethyl ester, gave the title compound (19 mg, 66%) as a white powder.

¹H-NMR (300 MHz, CDCl₃): δ 0.91-0.98 (m, 3H), 0.99-1.10 (m, 6H),1.11-1.38 (m, 4H), [(1.43 & 1.45) s, 9H], 1-45-1.94 (m, 9H), 2.05-2.18(m, 1H), 2.22-2.40 (m, 1H), 3.16-3.24 (m, 1H), 3.77 (s, 3H), 3.98 (s,3H), 4.04-4.18 (m, 1H), 4.36-4.57 (m, 3H), 6.00-4.08 (m, 1H), 6.13-6.21(m, 1H), 6.62-6.70 (m, 1H), 8.81-6.85 (m, 1H), 7.05-7.18 (m, 3H),7.41-7.57 (m, 4H), 8.02-8.13 (m, 3H). ¹³C-NMR (75.5 MHz, CDCl₃): δ 9.3,18.2, 19.0, [25.5 & 25.9], [28.0 & 28.3], 29.4, 31.4, 32.1, 35.7, 40.7,50.4, 52.2, 54.2, 55.5, 57.0, 58.2, 81.8, 82.4, 982, 107.5, 115.0,118.1, 122.9, 127.6, 128.7, 128.8, 128.9, 129.2, 135.1, 140.4, 142.2,151.4, 181.3, 163.9, 170.4, 170.9, 171.2, 172.0. MALDI-TOF m/z 821.60[(M+Na)⁺ calcd for C₄₅H₅₈N₄NaO₉ ⁺ 821.41].

Example 16

(S)-2-{[((3R,5R) &(3S,5S))-5-((S)-1-tert-Butoxycarbonyl-propylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarbonyl]-amino}-3-methyl-butyricacid methyl ester (16)

Reaction of 12 (24 mg, 44 mmol) according to the method described forthe preparation of 14 using D-valine methyl ester instead of(2-amino-3-methyl-butyrylamino)cyclohexyl acetic acid methyl ester, gavethe title compound (27 mg, 97%) as a white powder.

¹H-NMR (300 MHz, CDCl₃): δ 0.82-0.99 (m, 9H), [(1.42 & 1.44) s, 9H]1.65-1.95 (m, 2H), 2.18-2.25 (m, 1H), 2.26-2.40 (m, 1H), 3.20-3.25 (m,1H), 3.75 (s, 3H), 3.97 (s, 3H), 4.15-4.19 (m, 1H), 4.36-4.43 (m, 1H),4.64-4.75 (m, 1H), 6.03-6.15 (m, 1H), 6.80-6.85 (m, 2H), 7.10-7.20 (m,3H), 7.42-7.58 (m, 4H), 8.0-8.10 (m, 3H). ¹³C-NMR (75.5 MHz, CDCl₃): δ9.7, [18.2 & 19.1], 25.7, [28.1 & 28.2], 32.0, 35.6, 50.4, 52.4, 54.5,55.7, 57.6, 81.7, 82.7, 98.4, 107.7, 115.2, 118.4, 123.2, 127.8, 129.0,129.2, 129.5, 134.8, 135.0, 140.4, 142.5, 151.6, 159.6, [161.1 & 161.5],164.6, 171.1, 172.2. MALDI-TOF m/z 682.51[(M+Na)⁺ calcd for C₃₇H₄₅N₃NaO₈⁺ 682.31].

Example 17

(S)-2-{[(1R,4R) & (1S,4S))-2-{(S)-1-[(2,5-Dimethoxy-phenyl)-ethyl-carbamoyl]-2-methyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricacid tert-butyl ester (17)

Compound 17 (28.6 mg, 59%) was prepared from 12 (33 mg, 60 mmol)according to the method for the preparation of 14 using2-amino-N-(2,5-dimethoxy-phenyl)-N-ethyl-3-methyl butyramide instead of(2-amino-3-methyl-butyrylamino)-cyclohexyl acetic add methyl ester. Thisgave the title compound as a white powder.

¹H-NMR (300 MHz, CDCl₃): δ 0.75-0.95 (m, 9H) 1.05-1.18 (m, 3H), [(1.42 &1.44) s, 9H], 1.60-1.95 (m, 3H), 2.20-2.40 (m, 1H), 3.20-3.34 (m, 1H),3.60-3.80 (m, 2H), [3.62-3.65 (m, 3H)], [3.79-3.82 (m, 3H)], 3.98 (s,3H), 4.024-18 (m, 1H), 4.30-4.44 (m, 2H), 6.05-6.18 (m, 1H), 6.60-6.63(m, 1H), 6.77-6.80 (m, 2H), 6.85-6.93 (m, 2H), 7.12-7.20 (m, 2H),7.35-7.60 (m, 5H), 8.02-8.20 (m, 3H). ¹³C-NMR (75.5 MHz, CDCl₃): δ [9.6& 9.7], [12.5 & 12.8], [17.1 & 17.5], [19.4 & 19.5], 25.6, [28.0 &28.1], 32.4, 35.8, 43.0, 44.3, [50.2 & 50.3], 54.3, [54.8 & 55.0 & 55.2& 55.5], [55.6 & 55.7 & 55.9 & 56.0], 81.7, 82.8, 98.4, 106.9, [112.4 &112.5], 113.7, 115.0, 115.2, 115.9, 116.3, 118.4, [123.0 & 123.1],[127.7 & 127.8], 128.8, 128.9, 129.5, 130.1, [134.1 & 134.2], 142.6,149.1, 149.4, 153.4, 158.9, [161.4 & 161.6], [163.2 & 163.5], 170.9,[171.3 & 171.5], 172.3. MALDI-TOF m/z 831.62 [(M+Na)⁺ calcd forC₄₆H₅₆N₄NaO₉ ⁺ 831.39].

Example 18

(S)-2-{[((1R,4R) &(1S,4S))-2-{(S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricadd tert-butyl ester (18)

Compound 18 (16.1 mg, 26%) was prepared from 12 (43.2 mg, 0.077 mmol)according to the method for the preparation of 14 using(2-amino-3,3-dimethyl-butyrylamino)-cyclohexyl-acetic acid methyl esterinstead of (2-amino-3-methyl-butyrylamino)-cyclohexyl acetic acid methylester. Flash column chromatography was performed in toluene/ethylacetate 3:1 instead of 6:1: This gave the title compound as a whitepowder.

¹H-NMR (300 MHz, CDCl₃): δ 0.77-0.83 (m, 3H), [(0.92 & 0.93) s, 9H]0.94-1.20 (m, 4H), [(1.36 & 1.38) s, 9H], 1.42-1.76 (m, 8H), 2.20-2.38(m, 1H), 2.81-2.96 (m, 1H), 3.20-3.22 (m, 1H), 2.78 (s, 3H), [(3.83 &3.85) a, 3H], 3.97-4.02 (m, 1H), 4.17-4.21 (m, 1H), 4.22-4.37 (m, 2H),5.85-5.97 (m, 1H), [6.76-6.78 (m, 0.5H)], [6.80-6.82 (m, 0.5H)],6.98-7.05 (m, 3H), 7.23-7.41 (m, 8H), 7.82-7.99 (m, 3H). ¹³C-NMR (75.5MHz, CDCl₃): δ [9.4 & 9.5], [25.4 & 25.5], 25.8, [26.5 & 26.6], [27.9 &28.0], [28.4 & 28.51, 29.3, (35.4 & 35.7], [36.0 & 36.4], [40.5 & 40.7],[50.2 & 50.5), [52.1 & 52.2], 154.1 & 54.3], 55.5, [57.0 & 57.3], [60.4& 60.7], [81.8 & 82.0], [82.4 & 82.5] 98.1, 107.5, 115.0, 118.1, 123.0,127.5, 128.7, 128.8, 129.2, 134.9, 135.8, 141.9, 142.5, 151.3, 159A,[160.9 & 161.3], [163.7 & 163.9], [169.9 & 170.0] [170.0 & 171.3],[172.5 & 172.4]. MALDI-TOF m/z 835.68 [(M+Na)⁺ calcd for C₄₆H₆₀N₄NaO₉ ⁺835.43].

Example 19

(S)-2-{[(1R,4R)-2-{(S)-1-[((S)-Cyclohexyl-ethoxycarbonyl-methyl)-carbamoyl]-2-methy-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino)-pentanoicacid tert-butyl ester (19a) and (S)-2-{[(1S,4S)-2-((S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-pentanoicadd tert-butyl eater (19b)

The add 11 (0.051 g, 0.087 mmol) and(2-amino-3-methyl-butyrylamino)-cyclohexyl-acetic acid methyl ester(0.054 g, 0.21 mmol) were dissolved in DMF (1.5 mL) and cooled in anice-bath. DIPEA (16 mg, 0.12 mmol) and HATU (47 mg, 0.13 mmol) wereadded. After two and a half hours the mixture was co-concentrated withtoluene and methanol and then purified by flash column chromatography(toluene/ethyl acetate 3:1). Further purification was performed on HPLC(90% MeOH+0.2% TEA). This gave after co-concentration the twodiastereomers 19a (9.4 mg, 13%) and 19b (5.3 mg, 7%) as slightly yellowsyrups. After lyophilisation 19a and 19b were collected as whitepowders:

¹H-NMR (300 MHz, CDCl₃): δ 0.86-0.93 (m, 3H), 0.94-1.00 (m, 6H),1.00-1.41 (m, 7H), 1.46 (s, 9H), 1.50-1.88 (m, 8H), 2.05-2.20 (m, 1H),2.20-2.37 (m, 1H), 3.12-3.25 (m, 1H), 3.73 (s, 3H), 3.97 (s, 3H),4.05-4.20 (m, 1H), 4.40-4.55 (m, 3H), 6.02-6.18 (m, 1H), 6.30 (d, J=8.52Hz, 1H), 6.63 (s, 1H), 6.76 (d, J=8.51 Hz, 1H), 7.08-7.18 (m, 2H),7.42-7.56 (m, 5H), 8.00-8.12 (m, 3H); ¹³C-NMR (75.5 MHz, CD₃OD): δ 14.0,18.4, 19.3, 26.1, 28.3, 28.5, 29.7, 31.9, 34.9, 36.0, 41.0, 50.7, 52.4,53.3, 55.7, 57.2, 58.6, 82.0, 82.7, 98.4, 105.7, 107.7, 115.2, 118.4,123.2, 125.3, 127.9, 129.0, 129.1, 135.1, 138.0, 142.4, 151.6, 159.4,161.6, 164.3, 170.7, 171.2, 172.3. 19b: ¹H-NMR (300 MHz, CDCl₃): δ0.90-1.04 (m, 9H), 1.04-1.43 (m, 7H), 1.47 (s, 9H), 1.50-1.87 (m, 8H),2.10-2.27 (m, 1H), 2.33-2.45 (m, 1H), 3.10-3.20 (m, 1H), 3.73 (s, 3H),3.96 (s, 3H), 4.02-4.10 (m, 1H), 4.36-4.53 (m, 3H), 6.00-6.16 (m, 1H),6.30 (d, J=8.52 Hz, 1H), 6.73 (s, 1H), 6.86 (d, J=7.96 Hz, 1H),7.08-7.16 (m, 2H), 7.36-7.56 (m, 5H), 8.03-8.11 (m, 3H). ¹³C-NMR (75.5MHz, CD₃OD): δ 14.0, 18.6, 19.2, 26.1, 28.2, 28.7, 29.7, 34.5, 36.1,36.6, 40.8, 50.5, 52.4, 53.4, 55.7, 57.3, 59.1, 64.8, 82.3, 98.4, 105.8,107.8, 115.3, 118.4, 123.2, 127.8, 129.0, 129.4, 135.2, 1422, 144.9,151.0, 151.6, 159.2, 164.3, 164.3, 170.2, 171.6, 171.9

Example 20

(S)-2-{[(1R,4R)-2-((R)-1-[((S)-Cyclohexyl-methoxycarbonyl-methy)-carbamoyl]2,2-dimethyl-propylcarbamoyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-pentanoicacid tert-butyl ester (20a) and (S)-2-{[(1S,4S)-2-{(R)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-pentanoicacid tert-butyl ester (20b)

Method A:

The carboxylic acid 11 (57 mg, 0.10 mmol) was dissolved in warm (50° C.)dry THF (2 mL). (2-Amino-3,3-dimethyl-butyrylamino)-cyclohexyl-aceticadd methyl ester (50 mg, 0.12 mmol), DIPEA (30 mg, 0.23 mmol), DCC (25mg, 0.12 mmol) and HOBt (17 mg, 13 mmol) were added. After two hours themixture was concentrated and added to a short column (toluene/Ethylacetate 1:3+3% AcOH). Then it was further purified on HPLC using 90%MeOH+0.2% TEA. The diastereomeric products were not separated. AfterHPLC the solution was co-concentrated with toluene and methanol to give20 (28 mg, 34%).

Method B:

To an ice-cold solution of 11 (60 mg, 0.10 mmol) and(2-amino-3,3-dimethyl-butyrylamino)-cyclohexyl-acetic acid methyl ester(42 mg, 0.15 mmol) DIPEA (19 mg, 0.15 mmol) and HATU (62 mg, 0.16 mmol)were added. After two and a half hours the mixture was concentrated andpurified using column chromatography. (toluene/Ethyl acetate 3:1). Thediastereomeric mixture was separated using HPLC (90% MeOH+0.2% TEA).This gave 20a (6 mg, 6%) and 20b (9 mg, 10%).

20a: ¹H-NMR (300 MHz, CDCl₃): δ 0.82-0.90 (m, 3H), 1.01 (s, 9H),1.05-1.40 (m, 7H), 1.46 (s, 9H), 1.50-1.80 (m, 8H), 2.20-2.35 (m, 1H),3.07-3.25 (m, 1H), 3.73 (s, 3H), 3.97 (s, 3H), 4.11 (d, J=7.96 Hz, 1H),4.38-4.52 (m, 3H), 6.03-6.12 (m, 1H), 6.24 (d, J=8.79 Hz, 1H), 6.63 (s,1H), 6.82 (d, J=9.06 Hz, 1H), 7.07-7.27 (m, 2H), 7.36 (d, J=7.96 Hz,1H), 7.41-7.55 (m, 4H), 8.01-8.10 (m, 3H); ¹³C-NMR (75.5 MHz, CD₃OD): δ14.0, 18.8, 26.1, 26.8, 28.2, 28.6, 29.6, 34.9, 35.6, 36.2, 40.9, 50.7,52.4, 53.3, 55.7, 57.3, 60.8, 82.0, 82.7, 98.4, 105.2, 107.7, 115.2,118.4, 123.2, 127.9, 129.0, 129.4, 131.1, 135.1, 138.4, 142.4, 153.3,1569.6, 161.6, 164.2, 170.1, 171.3, 172.2. 20b: ¹H-NMR (300 MHz, CDCl₃):δ 0.90-0.98 (m, 3H), 1.04 (s, 9H), 1.08-1.40 (m, 7H), 1.44 (s, 9H),1.55-1.90 (m, 8H), 2.20-2.38 (m, 1H), 3.10-3.22 (m, 1H); 3.73 (s, 3H),3.97 (s, 3H), 4.02-4.15 (m, 1H), 4.35-4.48 (m, 3H), 6.00-6.08 (m, 1H),6.72 (s, 1H), 6.90 (d, J=9.06 Hz, 1H), 7.09-7.20 (m, 3H), 7.44-7.55 (m,5H), 8.03-8.11 (m, 3H).

Example 21

(1R,2S)-1-{[((1R,4R) & (1S,4S))-2-{(S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicacid tert-butyl ester (21)

The acid 13 (35 mg, 0.060 mmol) and(2-amino-3,3-dimethyl-butyrylamino)-cyclohexyl-acetic acid methyl ester(22 mg, 0.080 mmol) were dissolved in dry THF (1.5 mL) and warmed to 50°C. HOBt (11 mg, 0.080 mmol) and DCC (31 mg, 0.15 mmol) were added. Afterone hour the mixture was co-concentrated with toluene and methanol andthen purified by flash column chromatography (toluene/ethyl acetate1:1). Further purification was performed on HPLC (80% MeOH+0.2% TEA. Thediastereomeric mixture 21 was concentrated and gave a slightly yellowoil (26.4 mg, 53%). After lyophilisation 21 was collected as a whitepowder.

¹H-NMR (300 MHz, CDCl₃): δ [(0.98 & 1.00), s, 9H], 1.01-1.38 (m, 5H),[(1.39 & 1.40) s, 9H], 1.52-1.63 (m, 4H), 1.65-1.80 (m, 4H), 1.90-2.05(m, 1H), 2.20-2.40 (m, 1H), 3.02-3.20 (m, 1H), [(3.66 & 3.67) s, 3H),3.98 (s, 3H), 3.99-4.02 (m, 1H), 4.30-4.45 (m, 2H), 5.05-5.11 (m, 1H),5.20-5.30 (m, 1H), 5.60-5.81 (m, 1H), 6.03-6.17 (m, 1H), 6.77-6.82 (m,1H), 6.95-7.22 (m, 5H), 7.40-7.50 (m, 4H), 8.01-8.10 (m, 3H). ¹³C-NMR(75.5 MHz, CDCl₃): δ 22.3, [25.7 & 25.8], [26.4 & 26.5], [28.0 & 28.4]29.2, 32.7, 33.3, (35.3 & 35.4], 36.0, [40.2 & 40.3], 40.7, 52.0, 55.4,[57.2 & 57.4] [60.4 & 60.5], [87.6 & 87.7], [82.3 & 82.5], 98.4, 107.0,114.9, [117.4 & 117.5], 118.1, 122.9, 127.6, 128.8, 128.9, 129.2, [133.6& 133.8], 135.9, 136.9, 140.1, [141.4 & 141.6], 151.1, 159.6, [160.9 &161.3], [164.2 & 164.6], 168.9, 170.3, [172.1 & 172.6]. MALDI-TOF m/z859.77 [(M+Na)⁺ calcd for C₄₈H₆₀N₄NaO₉ ⁺ 859.43].

Example 22

(S)-2-{[(1R,4R)-2-{(R)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-pentanoicadd (22a) and (S)-2-{[(1S,4S)-2-{(R)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-pentanoicacid (22b)

The tert.butyl ester 20 (28 mg, 0.034 mmol), TES (8.7 mg, 0.075 mmol),DCM (1 mL) and TFA (1 mL) were mixed in a round bottomed flask. Twohours later the mixture was concentrated and the diastereomers wereseparated on HPLC using 65% MeOH+0.2% TEA as mobile phase. This gave 22a(15 mg, 65%) and 22b (12 mg, 45%) as slightly yellow syrups. Afterlyophilisation the title compounds were collected as white powders.

22a: [α]²²D+155.8; ¹H-NMR (300 MHz, CD₃OD): δ 0.90-0.97 (m, 3H), 1.03(s, 9H), 1.05-1.50 (m, 7H), 1.50-1.80 (m, 8H), 2.43-2.55 (m, 1H),2.77-2.90 (m, 1H), 3.68 (s, 3H), 3.96 (s, 3H), 4.20-4.30 (m, 2H),4.31-4.40 (m, 1H), 4.45-4.50 (m, 1H), 6.03-6.11 (m, 1H), 6.98 (s, 1H),7.12-7.19 (m, 1H), 7.36 (s, 1H), 7.41 (d, J=22 Hz, 1H), 7.50-7.60 (m,3H), 8.03-8.10 (m, 3H): ¹³C-NMR (75.5 MHz, CD₃OD): δ 13.1, 19.1, 26.1,28.7, 28.9, 29.5, 34.3, 34.8, 35.9, 40.1, 50.8, 51.2, 54.8, 55.0, 57.9,60.7, 83.5, 99.1, 106.0, 115.2, 118.2, 123.3, 127.8, 128.0, 128.7,128.8, 129.7, 135.2, 139.8, 143.7, 150.6, 160.1, 162.2, 165.2, 171.7,172.2, 173.4. 22b: [α]²²D −72.3; ¹H-NMR (300 MHz, CD₃OD): δ 0.90-0.97(m, 3H), 1.02 (s, 9H), 1.07-1.35 (m, 7H), 1.53-1.90 (m, 8H), 2.46-2.61(m, 1H), 2.76-2.88 (m, 1H), 3.69 (s, 3H), 3.96 (s, 3H), 4.15-4.35 (m,2H), 4.37-4.41 (m, 1H), 4.42-4.47 (m, 1H), 6.02-6.12 (m, 1H), 7.02 (s,1H), 7.16 (dd, J=2.47, 9.34 Hz, 1H), 7.32 (s, 1H), 7.40 (d, J=2.47 Hz,1H), 7.48-7.58 (m, 3H), 8.03-8.12 (m, 3H); ¹³C-NMR (75.5 MHz, CD₃OD): δ13.0, 18.8, 25.9, 26.0, 28.8, 29.4, 34.2, 34.8, 36.3, 39.9, 48.8, 50.5,51.1, 54.8, 57.9, 60.5, 82.8, 99.0, 106.0, 115.1, 118.2, 123.1, 127.8,127.9, 128.7, 129.0, 129.5, 136.7, 139.8, 142.8, 150.6, 160.1, 162.0,162.2, 164.7, 172.1, 173.5.

Example 23

(S)-2-{[(1R,4R)-2-{(R)-1-[((R)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2-methy-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricadd (23a) and (S)-2-{[(1S,4S)-2-{(R)-1-[((R)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricacid (23b)

Compound 23a (6.6 mg, 50%) and compound 23b (1.3 mg, 10%) were preparedfrom 15 (14 mg, 0.018 mmol) according to the method for the preparationof 22a and 22b. This gave the title compounds as white powders.

23a: ¹H-NMR (300 MHz, CD₃OD): 0.88-1.02 (m, 9H), 1.02-1.40 (m, 7H),1.55-1.97 (m, 6H), 2.01-2.10 (m, 1H), 2.38-2.52 (m, 1H), 2.88-3.00 (m,1H), 3.77 (s, 3H), 3.98 (s, 3H), 4.08-4.20 (m, 1H), 4.22-4.40 (m, 3H).6.03-6.18 (m, 1H), 6.86-6.99 (m, 1H), 7.08-7.20 (m, 1H), 7.23 (s, 1H),7.40-7.43 (m, 1H), 7.45-7.70 (m, 3H), 8.02-8.20 (m, 3H). ¹³C-NMR (75.5MHz, CD₃OD): δ 9.0, 17.6, 18.2, 24.5, 25.3, 28.1, 28.8, 30.9, 35.4,39.4, 49.6, 51.1, 54.7, 572, 58.0, 82.4, 98.5, 105.5, 114.5, 117.7,122.7, 127.2, 127.3, 128.2, 129.0, 135.6, 136.4, 141.7, 149.9, 159.5,161.2, 161.4, 164.0, 171.0, 171.7, 172.4. 23b: ¹H-NMR (300 MHz, CD₃OD):δ 0.9-1.20 (m, 9H), 1.21-1.53 (m, 7H), 1.55-1.93 (m, 6H), 2.05-2.20 (m,1H), 2.41-2.50 (m, 1H), 2.96-3-05 (m, 1H), 3.77 (s, 3H), 4.00 (s, 3H),4.05-4.40 (m, 4H), 6.05-6.18 (m, 1H), 6.90-6.95 (m, 1H), 7.05-7.22 (m,2H), 7.50-7.65 (m, 4H), 8.01-8.16 (m, 3H).

Example 24

(S)-2-{[((1R,4R) &(1S,4S))-2-{((S)-1-[((S)-Carboxy-cyclohexyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricacid (24)

The tert.butyl ester 14 (13.4 mg, 0.017 mmol), TES (4.83 mg, 0.042mmol), DCM (2 mL) and TFA (2 mL) were mixed n a round bottomed flask.One hour later the mixture was concentrated and purified by HPLC using65% MeOH+0.2% TEA as mobile phase. This gave 24 (4.3 mg, 34%) as aslightly yellow syrup. After lyophilisation 24 was collected as a whitepowder.

¹H-NMR (300 MHz, CD₃OD): δ 0.91-0.99 (m, 9H), 1.00-1.28 (m, 4H),1.65-1.78 (m, 9H), 1.92-1.95 (m, 1H), 2.00-2.05 (m, 1H), 2.93-3.01 (m,1H), 3.75 (s, 3H), 3.97 (s, 3H), 4.10-4.40 (m, 4H), 6.05-6.15 (m, 1H),6.88-6.94 (m, 1H), 7.05-7.10 (m, 2H), 7.41-7.43 (m, 1H), 7.44-7.55 (m,2H), 8.62-8.68 (m, 1H), 8.69-8.79 (m, 1H), 7.97-8.05 (m, 2H). ¹³C-NMR(75.5 MHz, CD₃OD): δ 9.2, 18.5, 25.5, [29.0 & 29.2], [30.0 & 30.5],35.3, 37.7, 39.7, 46.2, 50.0, [51.4 & 51.5], 53.6, 55.1, 57.1, 58.4,83.1, 98.9, 104.9, 114.6, 118.3, 123.0, 123.4, 127.5, 128.4, 128.5,129.7, 135.0, 142.1, 145.7, 1462, 159.2, 161.9, 164.3, 171.5, 171.9,172.2. MALDI-TOF m/z 791.27 [(M+K)⁺ calcd for C₄₂H₄₈KN₄O₉ ⁺ 791.31].

Example 25

(S)-2-{[((3R,5R) &(3S,5S))-5-((S)-1-Carboxy-propylcarbamoyl)-3-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-1-enecarbonyl]-amino}-3-methyl-butyricacid methyl ester (25)

Compound 25 (8.0 mg, 60%) was prepared from 18 (13.8 mg, 0.022 mmol)according to the method for the preparation of 24 which gave the titlecompound as a white powder.

¹H-NMR (300 MHz, CD₃OD): δ 0.83-1.02 (m, 9H), 1.88-1.80 (m, 1H),1.82-2.02 (m, 1H), 2.10-2.22 (m, 1H), 2.40-2.60 (m, 1H), 2.81-2.95 (m,1H), 3.75 (s, 3H), 4.00 (s, 3H), 4.18-4.22 (m, 1H), 4.27-4.40 (m, 2H),6.05-6.12 (m, 1H), 6.99-7.02 (m, 1H), 7.16-7.21 (m, 1H), 7.38 (s, 1H),7.40-7.43 (m, 1H), 7.48-7.61 (m, 3H), 7.98-8.12 (m, 3H).

Example 26

(S)-2-{[((1R,4R) &(1S,4S))-2-{(S)-1-[(2,5-Dimethoxy-phenyl)-ethyl-carbamoyl]-2-methyl-propylcarbamoyl}4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricadd (26)

Compound 26 (5.7 mg, 36%) was prepared from 17 (16.7 mg, 0.021 mmol)according to the method for the preparation of 24 which gave the titlecompound as a white powder.

¹H-NMR (300 MHz, CD₃OD): δ 0.75-0.81 (m, 6H), 0.82-0.98 (m, 3H),1.00-1.10 (m, 3H), 1.60-2.00 (m, 3H), 2.40-2.56 (m, 1H), 2.80-2.88 (m,1H), 3.18-3.24 (m, 1H), 3.40-3.48 (m, 1H), [3.67-3.80 (m, 6H)], 3.97 (s,3H), 4.10-4.20 (m, 1H), 4.21-4.40 (m, 2H), 6.02-8.17 (m, 1H), 6.75-6.82(m, 1H), 6.84-7.01 (m, 3H), 7.10-720 (m, 1H), 7.30-7.37 (m, 1H),7.40-7.43 (m, 1H), 7.50-7.60 (m, 3H), 8.00-8.17 (m, 3H). ¹³C-NMR (75.5MHz, CD₃OD): δ 9.6, [11.8 & 12.0], [17.2 & 17.4], 18.9, 25.0, 32.3,35.7, 43.3, 44.2, [50.3 & 50.5], [54.5 & 54.8 & 54.9 & 55.0], [55.1 &55.2 & 55.3 & 58.0], 58.7, 83.6, 99.3, 105.5, [112.5 & 112.7], 114.3,[15.1 & 115.2], 115.7, 116.1, 118.4, [123.3 & 123.4], 125.2, [128.0 &128.1, 128.8, 129.1, 129.8, [135.1 & 135.3], 139.2, (143.3 & 144.4],149.2, [149.6 & 149.9], 153.8, 159.9, 162.4, [163.9 & 164.5], 172.1,172.8, [173.6 & 173.7]. MALDI-TOF m/z 775.30 [(M+Na)⁺ calcd forC₄₂H₄₈N₄NaO₉ ⁺ 775.33].

Example 27

(S)-2-{[((R,4R) &(1S,4S))-2-{(S-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-butyricacid (27)

Compound 27 (6.0 mg, 72%) was prepared from 18 (8.6 mg, 0.011 mmol)according to the method for the preparation of 24. Purification by HPLC(60% methanol+0.2% TEA) gave the title compound as a white powder.

¹H-NMR (300 MHz, CD₃OD): δ 0.88-0.95 (m, 3H), 0.98 (s, 9H), 0.97-1.24(m, 4H), 1.57-1.62 (m, 3H), 1.58-1.78 (m, 4H), 1.79-1.99 (m, 1H),2.35-2.44 (m, 2H), 2.85-2.98 (m, 1H), [(3.67 & 3.69) 8, 3H], 3.94 (s,3H), 4.10-4.20 (m, 1H), 4.30-4.40 (m, 3H), 6.00-6.09 (m, 1H), [6.80-6.82(m, 0.5H)] [6.85-6.87 (m, 0.5H)], 7.05-7.19 (m, 2H), 7.38-7.55 (m, 4H),7.95-8.07 (m, 3H). ¹³C-NMR (75.5 MHz, CD₃OD): δ [9.1 & 9.2], [24.7 &24.9], [25.4 & 25.5], [25.9 & 26.0], [28.3 & 28.4], 28.9, [34.8 & 34.9],[35.6 & 35.9], [39.6 & 39.7], [49.9 & 50.13, [51.4 & 51.2], [53.9 &54.0] 55.0. 157.2 & 57.4], 60.0, [82.1 & 82.5], 98.6, 106.2, 114.7,117.8, 122.7, 127.5, 127.7, [128.4 & 128.5], 129.1, 135.3, 136.3, 141.6,142.0, 150.5, 159.8, [161.0 & 161.3] [164.0 & 164.1], [171.6 & 171.9],[1722 & 172.3], [173.0 & 173.2]. MALDI-TOF m/z 779.43 [(M+Na)⁺ calcd forC₄₂H₅₂N₄NaO₉ ⁺ 779.36].

Example 28

(S)-2-{[(1R,4R)-2-{(S)-1-{((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl}-2-methyl-propylcarbamoyl}-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-pentanoicacid tert-butyl ester (28)

The tert.butyl ester 19a (7.6 mg, 0.0094 mmol) and TES (2.4 mg, 0.021mmol) were dissolved in DCM (1 mL) and the mixture was cooled in anice-bath. TFA (1 mL) was added. After two hours the mixture wasconcentrated and purified on HPLC using 60% MeOH+0.2% TEA as mobilephase. This gave 28 (6.1 mg, 86%) as a slightly yellow syrup. Afterlyophilisation the title compound was collected as white powder. ¹H-NMR(300 MHz, CD₃OD+CDCl3 (1:1)): δ 0.90-1.00 (m, 9H), 1.00-1.30 (m, 7H),1.50-1.90 (m, 8H), 2.00-2.10 (m, 1H), 2.40-2.50 (m, 1H), 2.85-2.98 (m,1H), 3.65-3.72 (s, 3H), 3.99 (s, 3H), 4.15-4.22 (m, 1H), 4.24-4.35 (m,2H), 4.38-4.44 (m, 1H), 6.10-6.20 (m, 1H), 6.95-6.96 (m, 1H), 7.16-7.23(m, 1H), 7.31 (s, 1H), 7.42 (d, J=2.47 Hz, 1H), 7.53-7.72 (m, 3H),7.97-8.16 (m, 3H); ¹³C-NMR (75.5 MHz, CD₃OD+CDCl₃ 1:1): δ 13.5, 18.3,19.0, 26.0, 29.0, 29.7, 31.0, 34.1, 35.8, 40.2, 51.9, 55.9, 57.7, 58.9,63.5, 68.4, 84.0, 99.6, 104.8, 105.7, 115.1, 119.0, 123.7, 128.1, 128.9,129.1, 130.4, 131.3, 135.3, 138.0, 142.9, 159.5, 162.8, 164.8, 172.2,172.2, 172.4

Example 29

(S)-2-{[(1S,4S)-2-{(S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-pentanoicacid tert-butyl ester (29)

Compound 29 (1.3 mg, 28%) was prepared from 19b (5.3 mg, 0.065 mmol)according to the method for the preparation of 28. This gave the titlecompound as a white powder.

¹H-NMR (300 MHz, CD₃OD): δ 0.85-1.00 (m, 9H), 1.00-1.23 (m, 7H),1.50-1.78 (m, 8H), 2.05-2.23 (m, 1H), 2.50-2.66 (m, 1H), 2.70-2.85 (m,1H), 3.69 (s, 3H), 3.92 (s, 3H), 4.02-4.16 (m, 1H), 4.20-4.25 (m, 1H),4.35-4.40 (m, 2H), 6.09 (m, 1H), 7.00 (s, 1H), 7.12-7.18 (dd, J=2.47,2.19 Hz, 1H), 7.30 (s, 1H), 7.40 (d, J=2.42 Hz, 1H), 7.48-7.74 (m, 3H),8.03-8.10 (m, 3H); ¹³C-NMR (75.5 MHz, CDCl₃): δ 11.7, 16.5, 17.0, 24.4,27.2, 27.9, 29.0, 29.1 37.5, 41.8, 49.7, 50.5, 53.3, 56.3, 63.5, 66.5,81.0, 100.3, 101.0, 105.7, 113.6, 121.6, 128.3, 127.1, 127.9, 130.1,131.4, 135.6, 138.7, 141.1, 150.4, 160.2, 160.5, 165.3, 173.0, 173.6,173.7

Example 30

(1R,2S)-1-{[(1R,4R)-2-{(S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicacid (30a) and 1R,2S)-1-{[(1S,4S)-2-{(S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopent-2-enecarbonyl]-amino}-2-vinyl-cyclopropane-carboxylicacid (30b)

Compound 30a (6.3 mg, 49%) and compound 30b (5.6 mg, 43%) weresynthesized from 21 (13.8 mg, 0.0016 mmol) according to the method ofthe preparation of 22a and 22b. 30a and 30b: White powder.

30a: ¹H-NMR (300 MHz, CD₃OD): δ 1.02 (s, 9H), 1.03-1.43 (m, 5H),1.61-1.95 (m, 8H), 2.11-2.21 (m, 1H), 2.43-2.58 (m, 1H), 2.97-3.04 (m,1H), 3.78 (s, 3H), 4.01 (s, 3H), 4.02-4.17 (m, 1H), 4.25-4.40 (m, 2H),5.10-5-20 (m, 1H), 5.27-5.40 (m, 1H), 6.77-6.94 (m, 1H), 6.10-6.20 (m,1H), 6.97 (s, 1H), 7.18 (dd, J=2.5, 92 Hz, 1H), 7.22 (s, 1H), 7.46 (d,J=2.5 Hz, 1H), 7.52-7.65 (m, 3H), 8.00-8.18 (m, 3H). ¹³C-NMR (75.5 MHz,CD₃OD): δ 13.5, 25.3, 25.7, 28.3, 28.7, 29.0, 32.8, 34.6, 35.3, 39.3,49.7, 51.1, 54.6, 57.2, 59.8, 82.1, 98.4, 105.8, 114.5, 116.3, 117.6,122.6, 127.2, 128.1, 128.2, 128.8, 130.2, 133.7, 136.0, 139.5, 141.5,150.3, 159.7, 161.0, 1612, 163.4, 171.6, 172.5. MALDI-TOF m/z 803.56[(M+Na)⁺ calcd for C₄₄H₅₂N₄NaO₉ ⁺ 803.36]. 30b: ¹H-NMR (300 MHz, CD₃OD):δ 1.03 (s, 9H), 1.04-1.42 (m, 5H), 2.60-2.90 (m, 8H), 2.17-2.22 (m, 1H),2.40-2.55 (m, 1H), 2.96-3.10 (m, 1H), 3.77 (s, 3H), 4.01 (s, 3H),4.05-4.16 (m, 1H), 4.30-4.40 (m, 2H), 5.15-5.20 (m, 1H), 5.25-5.40 (m,1H), 5.78-5.95 (m, 1H), 6.10-6.20 (m, 1H), 6.98 (s, 1H), 7.17 (dd,J=2.5, 9.1 Hz, 1H), 7.28 (s, 1H), 7.46 (d, J=2.5 Hz, 1H), 7.50-7.65 (m,3H), 8.03-8.28 (m, 3H). ¹³C-NMR (75.5 MHz, CD₃OD): δ 13.7, 26.0, 26.3,28.8, 29.4, 29.6, 34.0, 35.2, 35.8, 40.1, 50.6, 51.7, 55.3, 57.8, 80.6,83.0, 99.1, 106.3, 115.2, 117.0, 118.3, 1232, 127.9, 128.0, 128.8,129.6, 130.6, 134.4, 136.1, 140.0, 142.5, 150.8, 160.3, 161.8, 162.0,165.7, 172.3, 173.0

Example 31

trans-(3R,4R)-Bis(methoxycarbonyl)cyclopentanol (31)

Sodium borohydride (1.11 g, 0.029 mol) was added to a stirred solutionof (1R,2S)-4-oxo-cyclopentane 1,2-dicarboxylic add dimethyl ester (4.88g, 0.0244 mol) in methanol (300 mL) at 0° C. After 1 h the reaction wasquenched with 90 mL brine, concentrated and extracted with ethylacetate. The organic phases were pooled, dried, filtered andconcentrated. The crude product was purified by flash columnchromatography (toluene/ethyl acetate 1:1) to give 31 (3.73 g, 76%) as ayellow oil.

Example 32

3-Oxo-2-oxa-bicyclo[22.2.1]heptane-6-carboxylic acid (32)

Sodium hydroxide (1M, 74 ml, 0.074 mol) was added to a stirred solutionof 31 (3.73 g, 0.018 mol) in methanol (105 mL) at room temperature.After 4 h, the reaction mixture was neutralized with 3M HCl, evaporatedand co-evaporated with toluene several times. Pyridine (75 mL) and Ac₂O(53 mL) were added and the reaction mixture was allowed to shakeovernight at room temperature. The mixture was then co-evaporated withtoluene and purified by flash column chromatography (ethyl acetate+1%acetic acid) to give 32 (2.51 g, 88%) as a yellow oil.

Example 33

3-Ox-2-oxa-bicyclo[2.2.1]heptane-5-carboxylic acid tert-butyl ester (33)

DMAP (14 mg, 0.115 mmol) and Boc₂O (252 mg, 1.44 mmol) was added to astirred solution of 32 (180 mg, 1.15 mmol) in 2 mL CH₂Cl₂ under inertargon atmosphere at 0° C. The reaction was allowed to warm to roomtemperature and was stirred overnight. The reaction mixture wasconcentrated and the crude product was purified by flash columnchromatography (toluene/ethyl acetate gradient 15:1, 9:1, 6:1, 4:1, 2:1)to give 33 (124 mg, 51%) as white crystals.

¹H-NMR (300 MHz, CD₃OD) δ 1.45 (s, 9H), 1.90 (d, J=11.0 Hz, 1H),2.10-2.19 (m, 3H), 2.76-2.83 (m, 1H), 3.10 (s, 1H), 4.99 (s, 1H);¹³C-NMR (75.5 MHz, CD₃OD) δ 27.1, 33.0, 37.7, 40.8, 48.1, 81.1, 81.6,172.0, 177.7.

Example 34

(1R,2R,4S)-2-((1R,2S)-1-Ethoxycarbonyl-2-vinyl-cyclopropylcarbamoyl)-4-hydroxy-cyclopentanecarboxylicacid tort-butyl ester (34)

Compound 33 (56 mg, 0.264 mmol) was dissolved in dioxane/water 1:1 (5mL) and the mixture was cooled to 0° C. 1 M lithium hydroxide (0.62 mL,0.520 mmol) was added and the mixture was stirred at 0° C. for 45minutes, after which the mixture was neutralized with 1M hydrochloricacid and evaporated and coevaporated with toluene. The residue wasdissolved in DMF (5 mL) and (1R,2S)-1-amino-2-vinylcyclopropanecarboxylic acid ethyl ester hydrochloride (60 mg, 0.313 mmol) anddiisopropylethylamine (DIEA) (138 □L, 0.792 mmol) were added and thesolution was cooled to 0° C. HATU (120 mg, 0.316 mmol) was added and themixture was stirred for 0.5 h at 0° C. and for an additional 2 h at roomtemperature. The mixture was then evaporated and extracted with EtOAc,washed with brine, dried, filtered and concentrated. Purification byflash column chromatography (toluene/EtOAc 1:1) provided compound 34 (86mg, 89%) as a colorless oil.

Example 35

(1R,2R,4R)-2-((1R,2S)-1-Ethoxycarbonyl-2-vinyl-cyclopropylcarbamoyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarboxylicacid tert-butyl ester (35)

Compound 34 (73 mg, 0.199 mmol) was dissolved in dry THF (4 mL) and2-phenyl-7-methoxy-4-quinolinol (88 mg, 0.342 mmol) andtriphenylphosphine (141 mg, 0.538 mmol) were added. The mixture wascooled to 0° C. and DIAD (0.567 mmol) dissolved in 1 mL THF was addeddropwise. The mixture was stirred for 48 h at room temperature. Thesolvent was evaporated and the crude product was purified by flashcolumn chromatography gradient elution (toluene/EtOAc 9:1, 6:1, 4:1) togive compound 35 (81 mg, 68%).

Example 36

Boc-L-ter-leucine-OH (38)

Triethylamine (890 uL, 6.40 mmol) was added dropwise to a stirredsolution of L-tert-leucine (300 mg, 2.29 mmol) and di-tert-butyldicarbonate (599 mg, 2.74 mmol) in dioxane/water 1:1 (8 mL) and thesolution was stirred overnight. The mixture was extracted with petroleumether (2×) and the aqueous phase was cooled to 0° C. and carefullyacidified to pH 3 by slow addition of 4M NaHSO₄.H₂O. The acidified waterphase was extracted with EtOAc (3×) and the combined organic phases werewashed with brine (2×) and was then dried, filtered and concentrated togive compound 36 (522 mg, 99%) as a colorless powder. No furtherpurification was needed. ¹H-NMR (300 MHz, CD₃OD) δ 0.99 (s, 9H), 1.44(s, 9H), 3.96 (s, 1H); ¹³C-NMR (75.5 MHz, CD₃OD) δ 27.1, 28.7, 34.9,68.0, 80.5, 157.8, 174.7.

Example 37

((S)-Cyclohexyl-methylcarbamoyl-methy)-carbamic acid tert-butyl ester(37)

Boc-Chg-OH (387 mg, 1.50 mmol) was coupled to methylamine hydrochloride(111 mg, 1.65 mmol) using the same HATU coupling conditions as in thesynthesis of compound 34. The crude product was extracted with EtOAc,washed with brine and concentrated. Purification by flash columnchromatography (EtOAc) provided compound 37 (307 mg, 76%) as a colorlesssolid.

¹H-NMR (300 MHz, CDCl₃) δ 0.91-1.13 (m, 2H), 1.14-1.31 (m, 3H), 1.44 (s,9H), 1.61-1.80 (m, 6H), 2.80 (d, J=4.7 Hz, 3H), 3.91 (dd, J=7.1, 9.1 Hz,1H), 5.23 (b, 1H), 6.52 (be, 1H); ¹³C-NMR (75.5 MHz, CDCl₃) δ 25.9,28.0, 26.1, 28.3, 28.5, 29.6, 40.5, 59.5, 79.7, 155.9, 172.4.

Example 38

{(S)-1-[((S)-Cyclohexyl-methylcarbamoyl-methyl)-carbamoyl]-2,2-dimethyl-propyl}-carbamicadd tert-butyl ester (38)

To a solution of compound 37 (98 mg, 0.362 mmol) in methylene chloride(3 mL) were added triethylsilane (115 mL, 0.742 mmol) and TFA (3 mL).The mixture was stirred for 2 h at room temperature and was thenevaporated and coevaporated with toluene. The deprotected amine wasdissolved in DMF (5 mL) and coupled to compound 36 (84 mg, 0.363 mmol)using the same HATU coupling conditions as in the synthesis of 34. Thecrude product was extracted with EtOAc, washed with brine, dried,filtered and concentrated. Purification by flash column chromatography(toluene/EtOAc 1:1) provided compound 38 (128 mg, 92%) as a colorlesssolid.

¹H-NMR (300 MHz, CDCl₃) δ 0.99 (s, 9H), 1.02-1.30 (m, 5H), 1.44 (s, 9H),1.58-1.77 (m, 4H), 1.78-1.89 (m, 2H), 2.79 (d, J=4.7 Hz, 3H), 4.11 (d,J=9.3 Hz, 1H), 4.33 (app. t, J, 8.5 Hz, 1H), 5.65 (b, 1H), 7.25 (b, 1H),7.39 (b, 1H); ¹³C-NMR (75.5 MHz, CDCl₃) δ 25.9, 25.9, 26.0, 26.2, 26.8,28.4, 29.0, 29.7, 34.5, 39.7, 58.4, 62.4, 79.4, 156.0, 171.4, 171.8.

Example 39

(1R,2S)-1-{[(1R,2R,4S)-2-{(S)-1-[((S)-Cyclohexyl-methylcarbamoyl-methyl)-carbamoyl]-2,2-diethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicacid ethyl ester (39)

To a solution of compound 35 (30 mg, 0.050 mmol) in methylene chloride(1.5 mL) were added triethylsilane (21 □L, 0.132 mmol) and TFA (1.5 ml).The mixture was stirred for 2 h at room temperature and was thenevaporated and coevaporated with toluene. The amine 38 (1.3 eq) wasdeprotected in the same manner as compound 35 and was then coupled todeprotected compound 35 using the same HATU coupling conditions as inthe synthesis of 34. The crude product was extracted with EtOAc, washedwith brine, dried, filtered and concentrated. Purification using HPLC(MeOH/water 9.1+0.2% triethylamine) provided compound 39 (30 mg, 74%) asa colorless sold.

¹H-NMR (300 MHz, CD₃OD) δ 0.81-1.14 (m, 4H), 0.99 (s, overlapped, 9H),1.21 (t, J 7.1 Hz, 3H), 1.35-1.51 (m, 4H), 1.52-1.65 (m, 3H), 1.66-1.72(m, 2H), 2.03-2.20 (m, 2H), 2.24-2.39 (m, 1H), 2.46-2.56 (m, 1H), 2.66(s, 3H), 2.72-2.85 (m, 1H), 3.39-3.48 (m, 2H), 3.90 (s, 3H), 4.03-4.15(m, 3H), 4.44 (s, 1H), 5.09 (dd, J=1.9, 10.3 Hz, 1H), 5.19-5.27 (m, 1H),5.25 (dd, overlapped, 1H), 5.79 (ddd, J=8.8, 10.3, 17.2 Hz, 1H), 8.99(s, 1H), 7.07 (dd, J=2.5, 9.1, Hz, 1H), 7.29 (d, J=2.5 Hz, 1H),7.43-7.52 (m, 3H), 7.86-7.98 (m, 2H), 8.05 (d, J=9.3 Hz, 1H); ¹³C-NMR(75.5 MHz, CD₃OD) δ14.7, 23.4, 28.0, 26.9, 27.1, 27.3, 30.1, 30.7, 35.0,35.4, 38.3, 38.8, 40.9, 41.0, 47.9, 55.9, 59.6, 62.0, 62.4, 79.8, 99.9,107.3, 116.4, 118.0, 119.1, 124.4, 128.9, 129.8, 130.5, 135.3, 141.3,152.1, 161.1, 162.4, 163.0, 171.6, 172.5, 173.7, 175.2, 176.8.Maldi-TOF-spectrum: (M+H)⁺ calcd: 810.4, found: 810.5; (M+Na)⁺ calcd:832.4, found: 832.4; (M+K)⁺ calcd: 848.5, found: 848.4.

Example 40

(1R,2S)-1-{[(1R,2R,4S)-2-{(S)-1-[((S)-Cyclohexyl-methylcarbamoyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicacid (40)

To a solution of compound 39 (20 mg, 0.025 mmol) in THF/MeOH/water 2:1:1(2 mL) at 0° C. was added 1M LiOH (175 uL, 0.175 mmol) and the solutionwas allowed to attain room temperature and was stirred for 48 h. Thesolution was acidified to pH 3 with 1M HCl and was then evaporated andcoevaporated with toluene. The crude product was purified by HPLC(MeOH/water 6:4+0.5% TFA followed by MeOH/water 4:1+0.2% TFA) to givecompound 40 (13 mg, 67%) as a colorless solid. ¹H-NMR (300 MHz, CD₃OD) δ0.82-0.98 (m, 1H), 1.01 (s, 9H), 1.05-1.26 (m, 3H), 1.34-1.43 (m, 1H),1.49-1.77 (m, 8H), 2.10-2.21 (m, 1H), 2.28-2.42 (m, 2H), 2.50-2.61 (m,1H), 2.64 (s, 3H), 2.68-2.81 (m, 1H), 3.36-3.45 (m, 2H), 4.04-4.11 (m,1H), 4.06 (s, overlapped, 3H), 4.27 (d, J=8.8 Hz, 1H), 5.10 (dd, J=1.8,10.3 Hz, 1H), 5.28 (dd, J=1.8, 17.2 Hz, 1H), 5.59-5.68 (m, 1H), 5.82(ddd, J=9.1, 10.3, 17.2 Hz, 1H), 7.44 (dd, J=2.5, 11.8 Hz, 1H), 7.50 (s,1H), 7.53 (d, J=2.5 Hz, 1H), 7.69-7.78 (m, 3H), 8.02-8.07 (m, 2H), 8.39(d, J=9.3 Hz, 1H); ¹³C-NMR (75.5 MHz, CD₃OD) δ 23.5, 26.0, 26.9, 27.2,27.3, 30.0, 30.7, 34.7, 35.3, 37.0, 38.7, 41.0, 41.3, 47.4, 56.9, 59.4,62.7, 83.9, 100.4, 1022, 1162, 117.7, 121.7, 126.7, 129.8, 130.8, 133.4,133.9, 135.6, 143.5, 158.0, 166.6, 168.6, 172.5, 173.4, 173.6, 175.4,176.4. Maldi-TOF-spectrum: (M+H)⁺ calcd: 782.4, found: 782.2; (M+Na)⁺calcd: 804.4, found: 804.2; (M+K)⁺ calcd: 820.5, found: 820.2.

Example 41

3-Oxo-2-oxa-bicyclo[2.2.1]heptane-5-carboxylic acid meth ester (41)

Compound 32 (1.014 g, 6.50 mmol) was dissolved in acetone (35 mL) beforemethyl Iodide (13.68 g, 96.4 mmol) and sliver(I)oxide (1.61 g, 6.95mmol) were added. After stirring for 3 h the mixture was filteredthrough celite and the filtrate was evaporated before purification byflash column chromatography (toluene/ethyl acetate 4:1) was performedyielding the methyl ester 41 (702 mg, 64%) as white crystals. ¹H-NMR(300 MHz, CDCl₃): δ 1.96 (d, J=10.7 Hz, 1H), 2.21-2.25 (m, 3H),2.91-2.95 (m, 1H), 3.16 (s, 1H), 3.75 (s, 3H), 4.98 (app. s, 1H).

Example 42

(1R,2R,4S)-2-((S)-1-tert-Butoxycarbonyl-butylcarbamoyl)-4-hydroxy-cyclopentanecarboxylicacid methyl ester (42)

Compound 41 (263 mg, 1.55 mmol) and H-Nva-OtBu (420 mg, 2.42 mmol) weredissolved in dry THF (20 mL). DIEA (530 uL, 3.04 mmol) and2-hydroxypyridine (260 mg, 2.73 mmol) were added and the mixture wasrefluxed for five days. The solvent was evaporated and the crude productwas purified by flash column chromatography (toluene/EtOAc 1:2) to give42 (510 mg, 96%).

Example 43

(1R,2R,4R)-2-((S)-1-tert-Butoxycarbonyl-butylcarbamoyl)-4-(7-methoxy-2-phenylquinolin-4-yloxy)-cyclopentanecarboxylic acid methyl eater (43)

Compound 42 (249 mg, 0.725 mmol), 2-phenyl-7-methoxy-4-quinolinol (310mg, 1.23 mmol) and PPh₃ (580 mg, 2.21 mmol) were dissolved in dry THFand the temperature was lowered to 0° C. DIAD (435 uL 2.21 mmol)dissolved in 2 mL dry THF, was added to the mixture during five minutes.After two hours the temperature was raised to room temperature and thesolution was stirred overnight. Evaporation and purification by flashcolumn chromatography (toluene/EtOAc gradient 6:1 to 4:1) gave 43 (324mg, 78%).

Example 44

(S)-2-{[(1R,2R,4S)-2-{(S)-1-[((S)-Cyclohexyl-methylcarbamoyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-pentanoicacid tert-butyl ester (44)

Compound 43 (38 mg, 0.066 mmol) was dissolved in dioxane/water 1:1 (4mL) and the solution was cooled to 0° C. and 1 M LiOH (132 ul, 0.132mmol) was added. The temperature was raised to room temperature and thesolution was stirred for 2 hours after which it was neutralized byaddition of M HCl and evaporated and coevaporated with toluene. Theresidue and deprotected amine 38 (1.1 eq) was dissolved in DMF andcoupled using the standard HATU coupling conditions as in the synthesisof compound 34. The crude product was extracted with EtOAc, washed withbrine, dried, filtered and concentrated. Purification with HPLC(MeOH/water 9:1+0.2% TEA) provided compound 44 (44 mg, 81%) as acolorless solid.

¹H-NMR (CDCl₃, 300 MHz) rotamers (5:1) δ 0.79 (t, J=7.3 Hz, 3H),0.85-1.19 (m, 3H), 0.93 (s, overlapped, 9H), 1.20-1.35 (m, 2H), 1.39 (s,1.5H), 1.43 (s, 7.5H), 1.54-1.79 (m, 6H), 2.06-2.28 (m, 3H), 2.39-2.51(m, 2H), 2.66-2.78 (m, 1H), 2.74 (d, overlapped, J=4.7 Hz, 3H),3.42-3.68 (m, 2H), 3.84 (s, 2.5H), 3.88 (s, 0.5H), 4.19 (t, J=8.9 Hz,1H), 4.39-4.59 (m, 1H), 4.68 (d, J=9.6 Hz, 1H), 5.04-5.14 (m, 1H), 6.77(s, 1H), 6.88-7.06 (m, 2H), 7.26-7.47 (m, 6H), 7.53 (b, 1H), 7.85-7.97(m, 3H); ¹³C-NMR (75.5 MHz, CDCl₃) δ 13.7, 18.7, 25.6, 25.7, 26.0, 26.7,28.0, 28.9, 29.7, 34.5, 34.7, 37.7, 38.0, 39.2, 46.6, 47.7, 52.7, 55.3,58.5, 60.3, 77.9, 81.7, 98.0, 107.4, 115.0, 117.9, 122.8, 127.4, 128.6,129.0, 140.2, 151.2, 158.9, 160.6, 161.1, 170.9, 171.6, 171.8, 172.7,173.3. Maldi-TOF-spectrum: (M+H)⁺ calcd: 828.5, found: 828.6; (M+Na)⁺calcd: 850.5, found: 850.6; (M+K)⁺ calcd: 866.6, found: 866.6.

Example 45

(S)-2-{[(1R,2R,4S)-2-{(S)-1-[((S)-Cyclohexyl-methylcarbamoyl-methyl)-carbamoyl]-2,2-dimethyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-pentanoicacid (45)

Compound 44 (21 mg, 0.025 mmol) was dissolved in CH₂Cl₂ (1.5 mL) andtriethylsilane (10 uL, 0.063 mmol) and TFA (1.5 mL) were added. Thesolution was stirred for 2 hours at room temperature after which thesolvents were evaporated and co-evaporated with toluene to providecompound 45 (20 mg, 100%) as a colorless solid.

¹H-NMR (300 MHz, CD₃OD) δ 0.93 (t, overlapped, 3H), 0.98 (s, 9H),0.99-1.25 (m, 4H), 1.30-1.49 (m, 3H), 1.50-1.90 (m, 8H), 2.25-2.39 (m,2H), 2.54-2.62 (m, 1H), 2.64 (s, 3H), 2.72-2.87 (m, 1H), 3.34-3.57 (m,3H), 4.02-4.13 (m, 1H), 4.06 (s, overlapped, 3H), 427-4.36 (m, 1H),4.37-4.47 (m, 1H), 5.57-5.66 (m, 1H), 7.45 (dd, J=2.3, 9.2 Hz, 1H), 7.48(s, 1H), 7.54 (d, J=2.2 Hz, 1H), 7.69-7.79 (m, 3H), 8.01-8.07 (m, 2H),8.42 (d, J=9.3 Hz, 1H); ¹³C-NMR (75.5 MHz, CD₃OD) δ 14.0, 20.2, 26.0,26.9, 27.2, 30.1, 30.7, 34.6, 35.3, 37.2, 39.1, 41.2, 47.7, 53.7, 56.9,59.4, 59.5, 62.5, 83.7, 100.4, 101.3, 102.2, 116.2, 121.7, 126.7, 129.8,130.8, 133.3, 133.9, 143.5, 157.9, 166.6, 168.5, 172.5, 173.6, 175.3,175.4, 175.5.

Maldi-TOF-spectrum: (M+H)⁺ calcd: 772.4, found: 772.6; (M+Na)⁺ calcd:794.4, found: 794.6; (M+K)⁺ calcd: 810.5, found: 810.6.

Example 46

Hept-6-enal (46)

To a solution of hept-6-en-1-ol (1 mL, 7.44 mmol) and N-methylmorpholineN-oxide (1.308 g, 11.17 mmol) in DCM (17 mL) was added ground molecularsieves (3.5 g, 4 Å). The mixture was stirred for 10 min at roomtemperature under nitrogen atmosphere before tetrapropylammoniumperruthenate (TPAP) (131 mg, 0.37 mmol) was added. After stirring foradditional 2.5 h the solution was filtered through celite. The solventwas then carefully evaporated and the remaining liquid was purified byflash column chromatography (DCM) to give the volatile aldehyde 46 (620mg, 74%) as an ol.

Example 47

N′-Hept-6-en-(E)-ylidene-hydrazinecarboxylic acid tert-butyl ester (47)

To a solution of 46 (68 mg, 0.610 mmol) and tart-butyl carbazate (81 mg,0.613 mmol) in MeOH (5 mL) was added ground molecular sieves (115 mg, 3Å). The mixture was stirred for 3 h after which it was filtered throughcelite and evaporated. The residue was dissolved in dry THF (3 mL) andAcOH (3 mL). NaBH₃CN (95 mg, 1.51 mmol) was added and the solution wasstirred over night. The reaction mixture was diluted with saturatedNaHCO₃ solution (6 mL) and EtOAc (6 mL). The organic phase was washedwith brine, saturated NaHCO₃, brine, dried over MgSO₄ and evaporated.The cyanoborane adduct was hydrolyzed by treatment with MeOH (3 mL) and2 M NaOH (1.9 mL). The mixture was stirred for 2 h and the MeOH wasevaporated. H₂O (5 mL) and DCM (5 mL) were added and the water phase wasextracted three times with DCM. The combined organic phases were driedand evaporated. Purification by flash column chromatography(toluene/ethyl acetate 9:1 with 1% triethylamine and toluene/ethylacetate 6:1 with 1% triethylamine) provided 47 (85 mg, 61%) as an oil.

Example 48

(1R,2)-1-{[(1R,2R,4R)-2-(N′-tert-Butoxycarbonyl-N-hept-6-enyl-hydrazinocarbonyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicacid ethyl ester (48)

Scaffold molecule 35 (135 mg, 0.225 mmol) and triethylsilane (71 μL,0.447 mmol) was dissolved in DCM (2 mL) after which trifluoroacetic acid(TFA) (2 mL) was added. The mixture was stirred for 2 h and thereafterco-evaporated with toluene in order to remove the TFA. The residue wasdissolved in DMF (3 mL) and 47 (60 mg, 0.263 mmol) and DIEA (118 μL,0.677 mmol) were added. The temperature was lowered to 0° C. and thecoupling reagent O-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (HATU) (94 mg, 0.247 mmol) was added. The coldsolution was allowed to stir for half an hour and then for additional 16h in room temperature. The solvent was removed by heating the reactionflask in a water bath under diminished pressure. The residue wasthereafter dissolved in ethyl acetate and the organic phase was washedthree times with brine, dried, filtered and evaporated. Purification byHPLC (MeOH/H₂O 90.10 with 02% triethylamine) gave 48 (140 mg, 82%) as anoil.

¹H-NMR (300 MHz, CDCl₃, 40° C.): δ 1.22 (t, J=7.1 Hz, 3H), 1.28-1.42 (m,6H), 1.46 (s, 9H), 1.52-1.62 (m, 2H), 1.82-1.91 (m, 1H), 1.96-2.16 (m,3H), 2.18-2.34 (m, 2H), 2.42-2.56 (m, 1H), 2.58-2.72 (m, 1H), 3.42 (app.bs, 3H), 3.66-3.84 (m, 1H), 3.92 (s, 3H), 4.15 (q, J=7.1 Hz, 2H),4.88-5.02 (m, 2H), 5.07-5.18 (m, 2H), 5.20-5.32 (m, 1H), 5.63-5.84 (m,2H), 6.62 (bs, 1H), 6.94 (s, 1H), 7.09 (dd, J=2.6, 9.2 Hz, 1H),7.36-7.51 (m, 4H), 7.99-8.10 (m, 3H); ¹³C-NMR (75.5 MHz, CDCl₃): δ 14.3,23.0, 26.4, 26.6, 28.3, 28.6, 33.2, 33.5, 35.6, 37.6, 40.6, 44.7, 47.1,48.6, 55.5, 61.5, 81.9, 98.4, 107.9, 114.5, 115.6, 118.1, 123.2, 127.6,128.3, 128.7, 129.1, 133.5, 138.7, 140.7, 151.5, 154.5, 159.2, 160.9,161.5, 170.5, 174.2, 176.3.

Example 49

(Z)-(1R,4R,6S,16R,18R)-14-tert-Butoxycarbonylamino-18-(7-methoxy-2-phenyl-quinolin-4-yloxy)-2,15-dioxo-3,14-diaza-tricyclo[14.3.0.0^(4,6)]nonadec-7-ene-4-carboxylicacid ethyl ester (49)

A solution of 48 (158 mg, 0.209 mmol) in dry DCM (25 mL) was bubbledwith argon for 5 min. To the stirred solution under argon atmosphere wasthen added a solution of Hoveyda-Grubbs catalyst 2^(nd) generation (11mg, 0.018 mmol) in dry DCM (5 mL). The mixture was stirred at refluxunder argon atmosphere for 16 h. The solvent was evaporated andpurification by HPLC (MeOH/H₂O 90:10 with 0.2% triethylamine) yielded 49(107 mg, 70%) as a colorless solid.

¹H-NMR (300 MHz, CD₃OD): δ 1.03-1.22 (m, 1H), 1.28 (t, J=7.1 Hz, 3H),1.32-1.44 (m, 4H), 1.49 (s, 9H), 1.55-1.73 (m, 2H), 1.81-1.91 (m, 1H),2.04-2.28 (m, 3H), 2.30-2.52 (m, 3H), 2.53-2.70 (m, 1H), 2.86-3.00 (m,1H), 3.34-3.44 (m, 1H), 3.48-3.62 (m, 1H), 3.95 (s, 3H), 4.19 (q, J=7.1Hz, 2H), 4.32-4.48 (m, 1H), 5.20-5.33 (m, 1H), 5.34 (bs, 1H), 5.58-5.70(m, 1H), 7.10 (s, 1H), 7.14 (dd, J=2.5, 9.1 Hz, 1H), 7.39 (d, J=2.5 Hz,1H), 7.45-7.55 (m, 3H), 8.00 (d, J=8.0 Hz, 2H), 8.17 (d, J=9.3 Hz, 1H);¹³C-NMR (75.5 MHz, CD₃OD): δ 14.6, 23.4, 27.5, 27.7, 28.0, 28.5, 30.7,36.1, 38.1, 42.5, 45.8, 56.0, 62.7, 79.9, 82.8, 100.2, 107.4, 116.8,119.1, 124.5, 126.5, 128.9, 129.8, 130.5, 135.8, 141.5, 152.2, 156.4,161.3, 162.5, 163.1, 171.9, 175.8, 179.0. MALDI-TOF-spectrum: (M+H)⁺calcd: 727.4, found: 727.5.

Example 50

(Z)-(1R,4R,6S,16R,18R)-14-tert-Butoxycarbonylamino-18-(7-methoxy-2-phenyl-quinolin-4-yloxy)-2,15-dioxo-3,14-diaza-tricyclo[14.3.0.0^(4,6)]nonadec-7-ene-4-carboxylicacid (50)

To a solution of 49 (27 mg, 0.037 mmol) in THF/MeOH/H₂O 2:1:1 (5 mL) wasadded 1 M LiOH (300 μL, 0.300 mmol). The solution was stirred for 24 hat room temperature and finally for one hour at reflux. Afteracidification to pH 3-4 with 1 M HCl and evaporation the residue waspurified by HPLC (MeOH/H₂O 80:20 and MeOH/H₂O 90:10) providing 50 (12mg, 46%) as a colorless sold.

¹H-NMR (300 MHz, CD₃OD): δ 1.06-1.24 (m, 1H), 126-1.42 (m, 3H), 1.48 (s,9H), 1.52-1.73 (m, 3H), 1.80-1.90 (m, 1H), 2.02-2.15 (m, 1H), 2.15-2.40(m, 4H), 2.43-2.54 (m, 1H), 2.54-2.68 (m, 1H), 2.88-3.00 (m, 1H),3.35-3.48 (m, 1H), 3.49-3.66 (m, 1H), 3.96 (s, 3H), 4.32-4.48 (m, 1H),5.25-5.42 (m, 2H), 5.56-5.88 (m, 1H), 7.14 (s, 1H), 7.17 (dd, J=2.5, 9.1Hz, 1H), 7.40 (d, J=2.2 Hz, 1H), 7.46-7.58 (m, 3H), 8.00 (d, J=8.0 Hz,2H), 8.19 (d, J=9.1 Hz, 1H); ¹³C-NMR (75.5 MHz, CD₃OD): δ 23.6, 26.8,27.8, 28.3, 28.5, 30.5, 35.8, 38.1, 43.0, 45.5, 56.0, 802, 82.7, 100.4,106.9, 116.6, 1192, 124.7, 127.4, 129.0, 129.8, 130.7, 134.8, 140.9,151.6, 156.5, 161.1, 163.0, 163.4, 173.8, 175.7, 179.3.

Example 51

((S)-1-Cyclopentylcarbamoyl-2,2-dimethyl-propyl)-carbamic acidtert-butyl ester (51)

To a cold solution of 36 (133 mg, 0.575 mmol), cyclopentylamine (64 μL,0.648 mmol) and DIEA (301 μL, 1.73 mmol) in DMF (3 mL) was added thecoupling reagent HATU (240 mg, 0.631 mmol). The mixture was stirred forhalf an hour and for additional two hours at room temperature. Thesolvent was removed by heating the reaction flask in a water bath underdiminished pressure and the residue was dissolved in ethyl acetate,after which the organic phase was washed three times with brine, dried,filtered and evaporated. Purification by flash column chromatography(toluene/ethyl acetate 4:1) provided 51 (140 mg, 82%) as colorlesscrystals.

¹H-NMR (300 MHz, CDCl₃): δ 0.95 (s, 9H), 1.28-1.48 (m, overlapped, 2H),1.40 (s, 9H), 1.49-1.71 (m, 4H), 1.86-2.01 (m, 2H), 3.76 (b, 1H),4.09-4.23 (m, 1H), 5.32 (b, 1H), 5.91 (b, 1H); ¹³C-NMR (75.6 MHz,CDCl₃): δ 23.6, 23.7, 26.5, 28.3, 32.6, 33.1, 34.5, 51.0, 62.2, 79.4,155.9, 170.3.

Example 52

(1R,2S)-1-{[(1R,2R,4S)-2-((S)-1-Cyclopentylcarbamoyl-2,2-dimethyl-propylcarbamoyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino)}-2-vinyl-cyclopropanecarboxylicacid ethyl ester (52)

Compound 51 (298 mg, 0.048 mmol) and 35 (16 mg, 0.054 mmol) wasdeprotected and coupled according to the method for the preparation of39. Purification by HPLC (MeOH/H₂O 90:10 with 0.2% triethylamine) gave52 (22 mg, 63%) as a colorless solid.

¹H-NMR (CDCl₃, 300 MHz): δ 0.97 (s, 9H), 1.21 (t, J=7.1 Hz, 3H),1.26-1.37 (m, 1H), 1.38-1.46 (m, 2H), 1.48-1.58 (m, 4H), 1.78-1.85 (m,1H), 1.86-2.02 (m, 3H), 2.03-2.19 (m, 1H), 2.28-2.40 (m, 2H), 2.41-2.54(m, 1H), 2.64-2.78 (m, 1H), 3.10-3.24 (m, 1H), 3.30-3.44 (m, 1H), 3.95(s, 3H), 4.04-4.21 (m, 3H), 5.12 (dd, J=1.7, 10.3 Hz, 1H), 5.14-5.22 (m,1H), 5.28 (dd, J=1.7, 17.0 Hz, 1H), 5.59 (b, 1H), 5.75 (ddd, J=8.8,10.3, 17.0 Hz, 1H), 6.66-8.82 (m, 2H), 6.99 (s, 1H), 7.09 (dd, J=2.5,9.1 Hz, 1H), 7.41-7.55 (m, 4H), 7.99-8.09 (m, 3H); ¹³C-NMR (75.5 MHz,CDCl₃): δ 14.3, 22.9, 23.6, 23.6, 26.7, 32.7, 332, 33.7, 34.8, 35.9,36.6, 40.2, 46.4, 47.5, 51.3, 55.5, 61.1, 61.4, 78.0, 98.4, 107.1,115.2, 117.9, 118.2, 123.1, 127.6, 128.8, 129.3, 133.5, 159.1, 161.4,169.4, 189.9, 173.1, 174.0. MALDI-TOF-spectrum: (M+H)⁺ calcd: 725.4,found: 725.6; (M+Na)⁺ calcd: 747.4, found: 747.6; (M+K)⁺ calcd: 763.3,found: 763.5.

Example 53

(1R,2S)-1-{[(1R,2R,4S)-2-((S)-1-Cyclopentylcarbamoyl-2,2-dimethyl-propylcarbamoyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicacid (53)

To a solution of 52 (14 mg, 0.019 mmol) in dioxane/HO 1:1: (4 mL) wasadded 1 M LiOH (115 μL, 0.115 mmol). The solution was stirred for 24 hat room temperature. Thereafter an additional portion of LiOH (75 uL,0.075 mmol) was added and the solution was stirred for another 24 h.After acidification to approximately pH 3 with 1 M HCl andco-evaporation with toluene the residue was purified by HPLC (MeOH/H₂O70:30 with 02% TFA) yielding 53 (8 mg, 60%) as a colorless solid.

¹H-NMR (300 MHz, CD₃OD): δ 0.98 (s, 9H), 1.28-1.48 (m, 3H), 1.49-1.76(m, 5H), 1.78-1.94 (m, 2H), 2.10-2.24 (m, 1H), 2.26-2.45 (m, 2H),2.50-2.62 (m, 1H), 2.66-2.79 (m, 1H), 3.35-3.48 (m, 2H), 3.94-4.03 (m,1H), 4.06 (s, 3H), 4.16-4.24 (m, 1H), 5.10 (dd, J=1.8, 10.3 Hz, 1H),5.29 (dd, J=1.8, 17.2 Hz, 1H), 5.62 (b, 1H), 5.82 (ddd, J=9.1, 10.3,17.2 Hz, 1H), 7.43 (dd, J=2.5, 9.3 Hz, 1H), 7.50 (s, 1H), 7.50-7.69 (dd,overlapped, 1H), 7.67-7.80 (m, 3H), 8.01-8.11 (m, 2H), 8.39 (d, J=9.3Hz, 1H); ¹³C-NMR (75.5 MHz, CD₃OD): δ 24.7, 24.7, 27.3, 33.1, 33.6,34.7, 35.4, 36.9, 38.7, 41.0, 47.4, 52.3, 56.9, 62.3, 83.9, 100.4,102.3, 116.2, 117.7, 121.6, 126.7, 129.8, 130.8, 133.4, 133.8, 135.6,143.5, 158.0, 166.5, 168.6, 171.9, 173.4, 175.2, 176.4.MALDI-TOF-spectrum: (M+H)⁺ calcd: 697.4, found: 897.3; (M+Na)⁺ calcd:718.7, found: 719.3; (M+K)⁺ calcd: 735.3, found: 735.3.

Example 54

(S)-tert-Butoxycarbonylamino-cyclohexyl-acetic add methyl ester (54)

To a solution of Boc-Chg-OH (53 mg, 0206 mmol) in acetone (3 mL) wereadded methyl iodide (195 μL, 3.1 mmol) and sliver (I) oxide (53 mg,0.229 mmol). The mixture was allowed to stir over night in a reactionflask that was covered with aluminium foil Thereafter the solution wasfiltered through celite and evaporated. Purification by flash columnchromatography (toluene/ethyl acetate 15:1) provided methyl ester 54 (56mg, 100%) as a colorless oil.

¹H-NMR (300 MHz, CDCl₃): δ 1.00-1.34 (m, 5H), 1.44 (s, 9H), 1.54-1.82(m, 6H), 3.73 (s, 3H), 4.20 (dd, J=2.8, 5.0 Hz, 1H), 5.05 (bs, 1H);¹³C-NMR (75.5 MHz, CDCl₃): δ 26.0, 28.2, 28.3, 29.5, 41.1, 52.0, 58.3,79.7, 155.6, 172.9.

Example 55

(S)—((S)-2-Benzyloxycarbonylamino-3-methy-butyrylamino)-cyclohexyl-aceticacid methyl ester (55)

Compound 54 (93 mg, 0.343 mmol) was deprotected and coupled to Z-Val-OH(95 mg, 0.378 mmol according to the method for the preparation of 39.Flash column chromatography (toluene/ethyl acetate 4:1) gave 55 (131 mg,94%) as a colorless solid.

¹H-NMR (300 MHz, CDCl₃): δ 0.92-1.30 (m, 11H), 1.54-1.88 (m, 6H),2.02-2.18 (m, 1H), 3.72 (s, 3H), 4.05-4.18 (m, 1H), 4.52 (dd, J=3.0, 5.5Hz, 1H), 5.12 (s, 2H), 5.49 (bs, 1H), 6.52 (bs, 1H), 7.34 (s, 5H);¹³C-NMR (75.5 MHz, CDCl₃): δ 17.8, 19.0, 25.8, 28.2, 29.3, 31.2, 40.5,51.9, 56.8, 60.0, 88.8, 127.7, 127.9, 128.1, 128.3, 136.2, 156.3, 171.3,172.2.

Example 56

(S)-2-{[(1R,2R,4S)-2-{(S)-1-[((S)-Cyclohexyl-ethoxycarbonyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}-4(7-methoxy-2-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-pentanolacid tert-butyl ester (56)

To a solution of 55 (40 mg, 0.099 mmol) in ethanol (95%) (7.5 mL) wasadded palladium on active carbon (10%, 40 mg) and the mixture washydrogenated under pressure at room temperature for 2 h. The mixture wasfiltered through celite and evaporated. Compound 43 (38 mg, 0.083 mmol)was dissolved in dioxane/H₂O 1:1 (3 mL) and the mixture was cooled to 0°C. before 1 M LiOH (140 μL, 0.140 mmol) was added to the stirredsolution. After 1 h the mixture was neutralized with 1 M hydrochloricacid and the solvent was evaporated and co-evaporated with toluene. Theresidue was coupled to deprotected 55 using the same HATU couplingconditions as in the synthesis of compound 48. Purification by HPLC(MeOH/H₂O 90:10 with 0.2% triethylamine) gave 56 (56 mg, 88%) as acolorless sold. ¹H-NMR (300 MHz, CDCl₃): δ 0.82-0.96 (m, 9H), 0.82-1.22(m, overlapped, 6H), 1.23-1.40 (m, 2H), 1.44 (s, 9H), 1.50-1.69 (m, 4H),1.71-1.87 (m, 2H), 1.95-2.06 (m, 1H), 2.07-2.22 (m, 1H), 2.28-2.54 (m,3H), 2.60-2.75 (m, 1H), 3.08-3.28 (m, 1H), 3.30-3.49 (m, 1H), 3.70 (s,3H), 3.94 (s, 3H), 4.28-4.38 (m, 1H), 4.41-4.57 (m, 2H), 5.17 (b, 1H),6.54-6.70 (m, 2H), 6.74 (b, 1H), 6.95 (s, 1H), 7.09 (dd, J=2.5, 9.1 Hz,1H), 7.39-7.55 (m, 5H), 7.98-8.10 (m, 3H); ¹³C-NMR (75.5 MHz, CDCl₃): δ13.7, 18.1, 18.6, 19.2, 25.9, 28.0, 28.2, 29.6, 30.7, 34.6, 36.5, 37.6,40.8, 47.4, 47.5, 52.1, 52.8, 55.5, 56.8, 58.9, 77.8, 82.0, 98.3, 107.5,115.3, 118.1, 123.1, 127.5, 128.7, 129.1, 140.5, 151.4, 159.2, 160.7,161.3, 171.0, 171.5, 172.3, 172.8, 173.0. MALDI-TOF-spectrum: (M+H)⁺calcd: 815.5, found: 815.7; (M+Na)⁺ calcd: 837.4, found: 837.6; (M+K)⁺calcd: 853.4, found: 853.6.

Example 57

(S)-2-{[(1R,2R,4S)-2-{(S)-1-[((S)-Cyclohexyl-methoxycarbonyl-methy)-carbamoyl]-2-methyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-pentanoicacid (57)

Tert.butyl ester 56 (28 mg, 0.034 mmol) and triethylsilane (14 μL, 0.088mmol) was dissolved in DCM (2 mL) after which trifluoroacetic acid (2mL) was added and the mixture was stirred for 2 h. Co-evaporation withtoluene gave 57 (26 mg, 100%) as a colorless solid.

¹H-NMR (300 MHz, CD₃OD): δ 0.86-1.00 (m, 9H), 1.01-1.24 (m, 4H),1.36-1.46 (m, 2H), 1.48-1.75 (m, 8H), 1.70-1.89 (m, overlapped, 1H),1.96-2.12 (m, 1H), 2.22-2.40 (m, overlapped, 2H), 2.49-2.64 (m, 1H),2.72-2.91 (m, 1H), 3.26-3.40 (m, overlapped, 1H), 3.50-3.68 (m,overlapped, 1H), 3.62 (s, 3H), 4.05 (s, 3H), 4.09-4.17 (m, 1H),4.17-4.25 (m, 1H), 4.35-4.45 (m, 1H), 5.82 (b, 1H), 7.44 (dd, J=2.2, 9.3Hz, 1H), 7.49 (s, 1H), 7.53 (d, J=2.2 Hz, 1H), 7.65-7.78 (m, 3H),7.98-8.06 (m, 2H), 8.41 (dd, J=2.8, 9.3 Hz, 1H); ¹³C-NMR (CD₃OD, 75.5MHz): δ 13.9, 18.8, 19.7, 20.2, 27.0, 29.7, 30.5, 31.8, 34.6, 37.7,38.9, 41.1, 47.8, 52.3, 53.6, 56.9, 58.8, 58.9, 60.3, 83.8, 100.4,102.2, 116.2, 121.6, 126.7, 129.8, 130.8, 133.3, 133.8, 143.5, 157.9,166.5, 168.5, 173.3, 173.9, 175.5, 175.5, 175.6. MALDI-TOF-epectrum:(M+H)⁺ calcd: 759.4, found: 759.7; (M+Na)⁺ calcd: 781.4, found: 781.7;(M+K)⁺ calcd: 797.4, found: 797.7.

Example 58

(S)-2-{[(1R,2R,4S)-2-{(S)-1[((S)Cyclohexyl-methoxycarbonyl-methyl)carbamoyl]-2-methyl-propylcarbamoyl}-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-butyricacid (58)

The procedure described in example 42 was followed but with the use ofL-2-amino-N-butyric acid tert.butyl ester instead of H-Nva-OtBu. Theafforded compound was then reacted as described in example 43 which gave(1R,2R,4R)-2-((S)-1-tert-butoxycarbonyl-propylcarbamoyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)cyclopentanecarboxylic acid methyl ester. Coupling of this compound with55 as described in example 56 followed by esterhydrolysis as describedin example 57 gave 58 as a colourless solid.

¹H-NMR (300 MHz, CD₃OD): δ 0.82-0.99 (m, 9H), 0.82-1.40 (m, overlapped,6H), 1.48-1.78 (m, 6H), 1.80-1.95 (m, 1H), 1.97-2.12 (m, 1H), 2.22-2.40(m, overlapped, 2H), 2.51-2.64 (m, 1H), 2.71-2.90 (m, 1H), 3.16-3.39 (m,overlapped, 1H), 3.49-3.59 (m, 1H), 3.63 (s, 3H), 3.95 (s, 3H),4.12-4.23 (m, 2H), 4.28-4.38 (m, 1H), 5.31 (b, 1H), 7.43 (dd, J=2.2, 9.3Hz, 1H), 7.47 (s, 1H), 7.51 (s, 1H), 7.66-7.89 (m, 3H), 7.99-8.07 (m,2H), 8.42 (d, J=9.1 Hz, 1H); ¹³C-NMR (75.5 MHz, CD₃OD): δ 10.7, 18.8,19.7, 25.8, 27.0, 27.0, 29.7, 30.5, 31.8, 37.7, 38.9, 41.2, 47.9, 52.3,55.3, 56.9, 58.8, 60.6, 83.6, 100.7, 102.2, 118.3, 121.5, 126.7, 129.8,130.8, 133.7, 133.8, 143.9, 158.2, 166.4, 168.3, 173.3, 173.8, 175.2,175.5, 175.6. MALDI-TOF-spectrum: (M+H)⁺ calcd: 745.4, found: 744.9;(M+Na)⁺ calcd: 767.4, found: 766.9; (M+K)⁺ calcd: 783.5, found: 782.9.

Example 59

(S)-2-{[(1R,2R,4S)-2-{((R)-1-[((R)-Cyclohexyl-methoxycarbonyl-methyl)-carbamoyl]-2-methyl-propylcarbamoyl}-4-(7ethoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-butyricacid (59)

The procedure described in example 54 was followed but with the use ofBoo-D-cyclohexyglycine Instead of Boc-L-cyclohexyglycine. The affordedcompound was then reacted as described in example 55 followed bycoupling with(1R,2R,4R)-2-((S)-1-tert-Butoxycarbonyl-pentylcarbamoyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarboxylicacid methyl ester as described in example 56. Removal of the ester groupas described in example 57 gave compound 59 as a colourless solid.¹H-NMR (CD₃OD, 300 MHz): δ 0.82-1.02 (m, 9H), 1.04-1.42 (m, 6H),1.52-1.80 (m, 6H), 1.80-1.95 (m, overlapped, 1H), 2.00-2.14 (m, 1H),2.29-2.46 (m, 2H), 2.51-2.65 (m, 1H), 2.68-2.84 (m, 1H), 3.24-3.39 (m,overlapped, 1H), 3.47-3.60 (m, 1H), 3.67 (s, 3H), 4.07 (s, 3H), 4.18-427(m, 2H), 4.28-4.38 (m, 11H), 5.64 (app. bs, 1H), 7.44 (d, J=2.3, 6.9 Hz,1H), 7.42 (s, 2H), 7.67-7.81 (m, 3H), 8.04 (d, J=7.8 Hz, 2H), 8.41 (d,J=9.1 Hz, 1H); ¹³C-NMR (CD₃OD, 75.5 MHz): δ 10.8, 18.5, 19.6, 25.7,27.1, 27.1, 30.1, 30.8, 31.9, 37.3, 38.2, 41.1, 47.8, 52.3, 55.4, 56.9,59.0, 59.1, 60.2, 83.8, 100.5, 102.2, 116.3, 121.6, 126.8, 1298, 130.8,133.6, 133.8, 143.7, 158.1, 166.5, 168.5, 173.4, 173.8, 175.4, 175.7,175.7. MALDI-TOF-spectrum: (M+H)⁺ calcd: 745.4, found: 745.4; (M+Na)⁺calcd: 767.4, found: 767.4; (M+K)⁺ calcd: 783.5, found: 783.3.

Example 60

Resin bound 2-tert.butoxycarbonylamino-3,3-dimethylbutyric acid (60)

To Argonaut resin PS-TFP (1.38 mmol/g, 10 g) and2-tert-butoxycarbonylamino-3,3-dimethyl-butyric acid (4.5 g, 20.7 mmol)was added dichloromethane (40 mL) and DMF (10 mL). To this mixture wasadded DMAP (1 g, 8.28 mmol) and then DIC (9.5 mL, 60.7 mmol). After 3hrs of agitation at RT the resin was filtered and washed successivelywith DMF, THF, DCM, THF, DCM and ether and then dried in a vacuum.

Example 61

[1-(2-Hydroxy-indan-1-ylcarbamoyl)-2,2-dimethyl-propyl]-carbamic acidtertbutyl ester (61)

To a portion of 60 (200 mg) in DCM aminoindanol (0.14 mmol) was added.The mixture was agitated for 2 hrs. The liquid was filtered of and theresin washed with 2×DCM. The combined liquids were combined andconcentrated to dryness to afford the title compound (20.5 mg, 0.055mmol) Purity>95% by HPLC. M+H⁺ 363.15. ¹³C NMR δ_(c) (100 MHz; CDCl₃;Me₄Si) 27.0, 28.5, 34.2, 39.8, 50.8, 57.9, 68.2, 73.7, 124.8, 125.6,127.4, 128.5, 140.4, 171.6. ¹H NMR δ_(H) (400 MHz; CDCl₃; Me₄Si) 1.07(9H, s, CCH₃), 1.44 (9H, s, OCCH₃), 2.93 (1H, dd, J_(gem) 16.4 Hz,J_(3,2) 2.3 Hz, CH₂), 3.15 (1H, dd, J_(gem) 16.4 Hz, J_(3,2)5.2 Hz,CH₂),

Example 62

2-Amino-N-(2-hydroxy-indan-1-yl)-3,3-dimethyl butyramide (62)

Compound 61 was kept in DCM-TFA 2:1 (2 mL) for 60 min at RT. Thesolution was co-evaporated with toluene to dryness.

Example 63

(2-tert-Butoxycarbonylamino-3,3-dimethyl-butyrylamino)-cyclohexyl-aceticacid methyl ester (63)

To a solution of 2-tert.butoxycarbonylamino-3,3-dimethyl butyric acid(500 mg, 2.16 mmol), Amino-cyclohexyl-acetic add methyl ester (444 mg,2.59 mmol) and HATU (2 g, 5.40 mmol) in DMF (20 mL) was addeddiisopropylethylamine (1.88 mL, 10.8 mmol). The solution was stirred for1 hrs at r.t. and diluted with dichloromethane (40 mL). This solutionwas washed with aqueous. NaHCO₃ (sat) and water (×2), dried andconcentrated. The product was >95% pure. M+H⁺385.4.

Example 64

{1-[(Cyclohexyl-methylcarbamoyl-methyl)-carbamoyl]-2,2-dimethyl-propyl}-carbamicacid tert-butyl ester (64)

To compound 63 in EtOH-THF 1:2 was added a large excess of methylamine(30% in water) and left at rt. for 2 weeks. The solution wasconcentrated to dryness and the residue subjected to a short silica gelcolumn eluted with 2% MeOH in dichloromethane to give a pure (>95%)product M+H⁺ 384.5.

Example 65

2-Amino-N-(cyclohexyl-methylcarbamoyl-methyl)-3,3-dimethyl-butyramide(65)

Compound 64 was kept in dichloromethane-trifuoroacetic acid 2:1 for 1 hat rt and concentrated to dryness. The residue was dried in a vacuum for16 hr. Reversed phase C18 HPLC showed >95% purity M+H⁺ 283.1.

Example 66

1-(2-Amino-4-methoxyphenyl)ethanone (66)

m-Anisidine (10.0 g, 82 mmol) was dissolved in CH₂Cl₂ (50 mL), and thesolution was cooled to −50° C. BCl₃(1 M in CH₂Cl₂, 82 mL, 82 mmol) wasadded slowly during 20 min, after which the mixture was stirred at −50°C. for 30 min, followed by sequential addition of AcCl (6.0 mL, 84 mmol)and AlCl₃ (11 g, 82 mmol). The mixture was stirred at −50° C. for 1 hand was then allowed to assume rt. Alter stirring at rt overnight, thesolution was heated at 40° C. for 4 h, after which the mixture waspoured over ice. The aqueous mixture was made alkaline with 10% NaOH(w/v) and extracted with EtOAc (4×200 mL). The combined organic phaseswere washed with brine, dried (MgSO₄), and evaporated to give a blacksolid, which was purified by flash column chromatography (ether/CH₂Cl₂20:80). The resulting solid was recrystallized from ether/hexane to givecompound 93 as shiny tan leaflets (5.6 g, 42%).

Example 67 N-(tert-Butyl)-N′-isopropylthiourea (67)

To a solution of tert-butylisothiocyanate (5.0 mL, 39 mmol) in CH₂Chi(200 mL) were added isopropylamine (4.0 mL, 47 mmol) anddisopropylethylamine (DIEA) (6.8 mL, 39 mmol), and the mixture wasstirred at rt for 2 h. The reaction mixture was diluted with EtOAc,washed with 10% citric acid (2×), saturated NaHCO₃ (2×), H₂O (2×), andbrine (1×). The organic layer was dried (MgSO₄) and evaporated to yieldthe tide compound (3.3 g, 52%) as a white solid which was used withoutfurther purification.

Example 68 N-Isopropylthiourea (68)

Compound 67 (3.3 g, 20 mmol) was dissolved in conc. HCl (45 mL) and thesolution was refluxed for 40 min. The mixture was allowed to cool to rtand then cooled in an ice bath and basified to pH 9.5 with sold andsaturated NaHCO₃, after which the product was extracted into EtOAc (3×).The combined organic phases were washed with H₂O (2×) and brine (1×),dried (MgSO₄), and evaporated to yield the crude title compound (2.1 g,90%) which was used without further purification.

Example 69

2-(Isopropylamino)-1,3-thiazole-4-carboxylic acid hydrobromide (69)

A suspension of compound 68 (2.1 g, 18 mmol) and 3-bromopyruvic acid(3.0 g, 18 mmol) in dioxane (180 mL) was heated to 80° C. Upon reaching80° C. the mixture became clear, and soon thereafter the product startedto precipitate as a white solid. After 2 h of heating, the reactionmixture was cooled to rt and the precipitate was filtered off andcollected. This yielded pure title product (4.4 g, 94%).

Example 70

N-(2-Acetyl-5-methoxyphenyl)-2-(isopropylamino)-1,3-thiazole-4-carboxamide(70)

A mixture of compound 69 (4.4 g, 16.5 mmol) and the aniline derivative66 (2.75 g, 16.5 mmol) in pyridine (140 mL) was cooled to −30° C. (uponcooling, the clear solution became partially a suspension). POCl₃ (3.3mL, 35 mmol) was added slowly over a 5 min period. The mixture wasstirred at −30° C. for 1 h, and was then allowed to assume rt. Afterstirring at rt for 1.5 h the reaction mixture was poured over ice, andthe pH was adjusted to about 9-10 using solid and saturated NaHCO₃. Thecrude product was extracted into CH₂Cl₂ (3×) and the combined organicphases were dried (MgSO₄) and evaporated. The crude dark-beige solid waspurified by flash column chromatography (hexane/EtOAc 55:45) to givecompound 70 (5.6 g, 76%) as a pale yellow solid.

Example 71

2-[2-(Isopropylamino)-1,3-thiazol-4-yl]-7-methoxyquinolin-4-ol (71)

A solution of tBuOK (2.42 g, 21 mmol) in anhydrous t.BuOH (40 mL) washeated to reflux Compound 70 (1.8 g, 5.4 mmol) was added portion-wiseover a 5 min period, and the dark red solution formed was stirred atreflux for an additional 20 min. The mixture was cooled to rt, and HCl(4 M in dioxane, 8.0 mL, 32 mmol) was added, after which the reactionmixture was concentrated under vacuum. In order to assure that all ofthe HCl and dioxane were removed, the crude product was re-dissolved inCH₂Cl₂ twice and thoroughly evaporated to obtain the slightly impure HClsalt of compound 71 (1.62 g) as a brown solid. The product was dissolvedin CH₂C₁ and washed with saturated NaHCO₃, after which the aqueous phasewas extracted several times with CH₂Cl₂. The combined organic phaseswere dried (MgSO₄) and evaporated to give compound 71 (1.38 g, 81%) as alight brown solid (>95% pure according to HPLC tests). ¹H-NMR (MeOH-d₄,400 MHz): δ 1.30 (d, J=8.0 Hz, 6H), 3.93 (s, 3H), 3.95-4.07 (m, 1H),6.73 (s, 1H), 8.99 (dd, J=2.4, 9.2 Hz, 1H), 7.26 (d, J=2.4 Hz, 1H), 7.37(s, 1H), 8.10 (d, J=9.2 Hz, 1H).

Example 72

(1R,4R,5R)—N-[(1S)-1-[[[(1S)-1-Cyclohexyl-2-(methylamino)-2-oxoethyl]amino]carbonyl]-2,2-dimethylpropyl]-3-oxo-2-oxabicyclo[2.2.1]heptane-5-carboxamide(72)

To a solution of compound 32 (53 mg, 0.34 mmol) in DMF (9 mL) was addedcompound 65 (80 mg, 0.28 mmol) and DIEA (290 □L, 1.66 mmol). Thesolution was cooled to 0° C. and HATU (127 mg, 0.33 mmol) was added.After stirring at 0° C. for 1 h and at it for 1 h the solvent wasevaporated, and the crude product was purified by flash columnchromatography (EtOAc/toluene 2:1) to give compound 72 (110 mg, 92%) asa white solid.

Example 73

(1R)-1-[[[1R,2R,4R)-2-[[[(1 S)-1-[[[(1S)-1-Cyclohexyl-2-(methylamino)-2-oxoethyl]amino]carbonyl]-2,2-dimethylpropyl]amino]carbonyl]-4-hydroxycyclopentyl]carbonyl]amino]-2-ethenyl-cyclopropanecarboxylicacid ethyl ester (73)

Compound 72 (60 mg, 0.14 mmol) was dissolved in dioxane (3.5 mL) and H₂O(2.5 mL) and the solution was cooled to 0° C. LiOH (1 M, 280 □L, 0.28mmol) was added dropwise during 5 min, after which the reaction mixturewas stirred at 0° C. for 40 min. The pH was adjusted to 7 using 1 M HCl,and the solvents were evaporated. The residue was suspended in DMF (5mL) and 1-amino-2-vinyl-cyclopropanecarboxylic acid ethyl ester (32 mg,0.17 mmol), and DIEA (146 □L, 0.84 mmol) were added. After cooling to 0°C. HATU (64 mg, 0.17 mmol) was added and the mixture was stirred at 0°C. for 1 h and at t for 1 h. The solvent was evaporated and the productwas purified using flash column chromatography (EtOAc/MeOH 9:1) to givecompound 73 (87 mg, 82%) as a white solid.

Example 74

tert-Butyl(1R,2R,4R)-2-[[[(1R)-1-(ethoxycarbonyl)-2-vinylcyclopropyl]amino]carbonyl]-4-[[2-[2-(Isopropylamino)-1,3-thiazol-4-yl]-7-methoxyquinolin-4-yl]oxy]cyclopentanecarboxylate(74)

The title compound was prepared according to the procedure described inexample 76 method A but with the use of compound 34 instead of compound73. (Note: 4 equivalents of Ph₃P and DIAD were used. Chromatographyeluent Toluene/EtOAc 1:1.)

Example 75

(1R,2R,4R)-2-[[[(1R)-1-(Ethoxycarbonyl)-2-vinylcyclopropyl]amino]carbonyl]4-[[2-[2-(isopropylamino)-1,3-thiazol-4-yl]-7-methoxyquinolin-4-yl]oxy]cyclopentanecarboxylicacid (75)

To a solution of compound 74 (20 mg, 30 umol) in CH₂Cl₂ (2 mL) was addedTFA (2 mL) and Et₃SiH (10 uL, 63 umol). After 2 h the volatiles wereevaporated and the product was used without any purification step.Compound 75:18 mg, quant as a white solid.

Example 76

(1R)-1-[[[(1R,2R,4S)-2-[[[(1S)-1-[[[(1S)-1-Cyclohexyl-2-(methylamino)-2-oxoethyl]amino]carbonyl]-2,2-dimethylpropyl]amino]carbonyl]-4-[[7-methoxy-2-[2-[(1-methylethyl)amino]-thiazolyl]-4-quinolinyl]oxy]cyclopentyl]carbonyl]amino]-2-ethenyl-cyclopropanecarboxylicacid ethyl ester (76)

Method A:

To a solution of compound 73 (59 mg, 0.10 mmol) in dry THF (4 mL) wasadded the quinoline 71 (49 mg, 0.16 mmol) and Ph₃P (65 mg, 0.25 mmol).After cooling to 0° C. DIAD (50 uL, 0.25 mmol) was added dropwise during5 min. The solution was stirred at 0° C. for 1 h and at it for 48 h. Thesolvent was evaporated and the remainder was purified using flash columnchromatography (CHCl₃/2 M NH₃ in MeOH 95:5) to give compound 76 (9 mg,10%) as a white solid.

Method B:

Compound 75 was coupled to compound 65 according to the procedure inexample 72 which gave the title compound (82%).

Example 77

(1R)-1-[[[(1R,2R,4S)-2-[[[(1S)-1-[[[(1S)-1-Cyclohexyl-2-(methylamino)-2-oxoethyl]amino]carbonyl]-2,2-dimethylpropyl]amino]carbonyl]-4-[[7methoxy-2-[2-[(1-methylethyl)amino]-4-thiazolyl]-4-quinolinyl]oxy]cyclopentyl]carbonyl]amino]-2-ethenyl-cyclopropanecarboxylicacid (77)

Compound 76 (8 mg, 9 μmol) was dissolved in a mixture of MeOH (150 μL)and THF (100 uL). A solution of LiOH (1 mg, 42 μmol) in H₂O (25 □L) wasadded and the mixture was stirred at 50° C. overnight. The solution wasneutralized with HOAc and evaporated. The residue was suspended in CH₂Cand washed with H₂O. The organic phase was evaporated to give the titlecompound (8 mg, quant.) as a white solid.

¹H-NMR (MeOH-d₄, 400 MHz) (mixture of rotamers): δ 0.60-1.33 (m, 21H),1.35-1.73 (m, 12H), 1.90-2.42 (m, 2H), 2.51-2.75 (m, 6H), 3.20-3.38 (m,1H), 3.85 (s, 3H), 3.95-4.28 (m, 1H), 4.91-5.02 (m, 1H), 5.12-5.23 (m,1H), 5.64-5.83 (m, 1H), 7.01-7.11 (m, 1H), 7.25-7.40 (m, 1H), 7.42-7.57(m, 1H), 7.85-8.08 (m, 1H).

Example 78

2-Amino-3,3-dimethyl-N-thiophen-2-yl-methyl-butyramide (78)

The title compound was prepared as described in example 61 but with theuse of thiophene-2-methylamine instead of aminoindanole followed byremoval of the Boc group as described in example 62.

Example 79

2-Amino-N-(6-hydroxy-4,5,6,7-tetrahydro-benzo[b]thiophen-5-yl)-3,3-dimethyl-butyramide(79)

The title compound was prepared as described in example 61 but with theuse of 2-amino-4,5,6,7-tetrahydro-benzo[b]thiophen-5-ol instead ofaminoindanole followed by removal of the Boc group as described inexample 62.

Example 80

2-Amino-N-(2-diethylamino-ethyl-3,3-dimethyl-butyramide (80)

The title compound was prepared as described in example 61 but with theuse of N,N-diethylethylenediamine instead of aminoindanole followed byremoval of the Boc group as described in example 62.

Example 81

2-Amino-N-[2-(2-methoxy-phenoxy)-ethyl]-3,3-dimethyl-butyramide (81)

The title compound was prepared as described in example 61 but with theuse of 2-methoxyphenoxyethylamine instead of aminoindanole followed byremoval of the Boc group as described in example 62.

Example 82

2-Amino-1-(3-hydroxy-pyrrolidin-1-yl)-3,3-dimethyl-butan-1-one (82)

The title compound was prepared as described in example 61 but with theuse of (R)-3-pyrrolidinone instead of aminoindanole followed by removalof the Boc group as described in example 62.

Example 83

2-Amino-N-(1,1-dioxo-tetrahydro-1-thiophen-3-yl)-3,3-dimethyl-butyramide(83)

The title compound was prepared as described in example 61 but with theuse of 2-methoxyphenoxyethylamine instead of aminoindanole followed byremoval of the Boc group as described in example 62.

Example 84

Carbamic acid, [(1 S)-1-[[(phenylsulfonyl)amino]carbonyl]butyl]-,phenylmethyl ester (84)

To a stirred solution of Z-Nva-OH (150 mg, 0.59 mmol) in THF (6 mL), CDI(400 mg, 2.4 mmol) was added. The slurry was agitated for 30 min at RTfollowed by the addition of DBU (200 uL, 1.3 mmol) and a solution ofbenzenesulfonamide (250 mg, 1.59 mmol) in THF (2 mL). The mixture wasstirred at 60° C. for 48 hrs followed by concentration to dryness. Theresidue was dissolved in MeOH and subjected to HPLC purification to givethe title compound (118.5 mg, 0.304 mmol). Purity>95% by HPLC. M−H⁺389.0, +Na 412.96.

Example 85

(2S)-2-Amino-N-(phenylsulphonyl)pentanamide (85)

Compound 84 was dissolved in MeOH (5 mL) followed by the addition ofPd/C and subjected to hydrogenation for 2 hrs. The slurry was filteredthrough celite, washed with MeOH and concentrated to dryness to give thetitle compound. Yield 100%. M+H⁺ 257.3.

Example 86

4-(7-Methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentane-12-dicarboxylicacid 1-{[1-(cyclohexylmethyl-carbamoyl)-2-methyl-propyl]-amide}2-[(1-phenylmethanesulfonylaminocarbonyl-2-vinyl-cyclopropyl)-amide](86)

N-(tert-Butoxycarbonyl)-L-valine was attached to Argonaut resin PS-TFPas described in example 60 followed by reaction withcyclohexanemethylamine as described in example 61 and removal of the Bocgroup as described in example 62. The afforded amine was used in acoupling reaction with compound 35 as described in example 39 followedby hydrolysis of the ethyl ester as described in example 40 which gave1-{[2-[1-(cyclohexylmethyl-carbamoyl)-2-methyl-propylcarbamoyl]-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicadd. The afforded add was then treated as described in example 94 butusing toluenesulphonamide instead of cyclopropylsulphonamide which gavethe title compound. Yield 6%. Purity>95% by HPLC. M+H⁺ 864.32.

Example 87

Acetic acid(1S,2R)-1-((2S)-2-amino-3,3-dimethyl-butyrylamino)-indan-2-yl ester (87)A solution of compound 61 (4 g) was kept in pyridine-acetic anhydride2:1 for 30 min. DCM was added and the solution was washed with citricacid (aq) and NaHCO₃ (aq). The organic layer was concentrated to drynesswhich gave the acetylated product>90% pure by HPLC. The affordedcompound was then kept in a solution of 30% TFA In DCM for 1.5 hrs andthen concentrated to dryness. Co-evaporation twice from toluene gave thetitle product>90% pure by HPLC.

Example 88

(2S)-Methanesulphonic acid 2-tert.butoxycarbonylamino-4-methyl-pentylester (88)

To a solution of ((S)-1-hydroxymethyl-3-methylbutyl)-carbamic acidtert-butyl ester (25 g, 115 mmol) in dichloromethane (500 mL) cooled byan ice-water bath was successively added disopropylethylamine (35.7 g,276 mmol) and methanesulphonyl chloride (15.81 g, 138 mmol). Theresulting solution was stirred over night during which time the mixturewas allowed to gradually warm up to ambient temperature. The mixture waswashed successively with water, 10% citric acid (aq), water andsaturated NaHCO₃ (aq), then dried with Na₂SO₄ and concentrated to abrown solid (32.6 g, 96%) which was used in the next reaction withoutfurther purification.

Example 89

ii) ((1 S)-1-Azidomethyl-3-methyl-butyl)carbamic acid tert.butyl ester(89)

The mesylate from example 88 (32.6 g, 110 mmol) was treated with sodiumazide (21.45 g, 330 mmol) in DMF at 80° C. for 24 hrs. The solvent wasevaporated, the residue was taken up in DCM, filtered and washed withsaturated NaHCO₃ (aq). The solution was dried with Na₂SO₄ andconcentrated to a brown oil which was purified by flash chromatographyusing a gradient of ethyl acetate and hexane to afford the titlecompound as a white solid (19.55 g, 73%).

Example 90

(1S)-1-Azidomethyl-3methyl-butylamine (90)

((1 S)-1-Azidomethyl-3-methyl-butyl)-carbamic acid tert-butyl ester(9.64 g, 39.78 mmol) was treated with TFA (30 mL) in DCM (150 mL) for 3hrs, the mixture was evaporated under reduced pressure and the residuewas dissolved in ethyl acetate and washed with aqueous 1 M K₂CO₃, driedwith Na₂SO₄ and concentrated to a yellow liquid (4.55 g, 80%).

Example 91

1-{[2-Hex-5-enylcarbamoyl-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino}-2-vinyl-cycloprpanecarboxylicacid ethyl ester (91)

The tert.butyl ester of compound 35 was removed by treatment withtriethylsilane as described in Example 39. The afforded add (724 mg,1.33 mmol), hex-5-enylamine hydrochloride (271 mg, 2 mmol) anddiisopropylethylamine (1.85 ml, 10.65 mmol) was dissolved in DMF (20 ml)and cooled to 0° C. After 30 min. HATU (608 mg, 1.6 mmol) was added andthe flask was removed from the ice-bath. The reaction was followed withLC-MS. After 3 h the reaction mixture was extracted between EtOAc (100ml) and aqueous sodium hydrogencarbonate (15 ml). The EtOAc-phase wasdried over magnesium sulphate, evaporated and purified by chromatographyon silica gel (25% EtOAc in hexane→50% EtOAc in hexane) to give the puretitle product (726 mg, 87%). MS (M+H⁺): 525.8

Example 92

17-(7-Methoxy-2-phenyl-quinolin-4-yloxy)-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carboxylicacid ethyl ester (92)

Compound 91 (363 mg, 0.58 mmol) was dissolved in degasseddichloromethane (100 ml). Hoveyda-Grubbs catalyst 2nd generation (26 mg,0.041 mmol) was added and the mixture was refluxed under argonatmosphere overnight. The reaction mixture was evaporated on silica andpurified by silica gel chromatography (50% EtOAc in hexane→70% EtOAc inhexane) to give the pure title product (111 mg, 32%). MS (M+H⁺): 597.7

Example 93

17-(7-Methoxy-2-phenyl-quinolin-4-yloxy)-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carboxylicadd (93)

Compound 92 (96 mg, 0.159 mmol) was dissolved in tetrahydrofuran (10ml), methanol (5 ml) and water (4 ml) Lithium hydroxide (40 mg, 1.67mmol) was dissolved in water (1 ml) and added. The reaction mixture washeated to 65° C. After 3 h the reaction mixture was cooled, acidifiedwith aqueous HCl (pH5), evaporated on silica and purified by silica gelchromatography (10% MeOH in dichloromethane→15% MeOH in dichloromethane)to give the pure title product (65 mg, 72%). MS (M+H⁺): 569.8

Example 94

Cyclopropanesulphonic add[17-(7-methoxy-2-phenyl-quinolin-yloxy)-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*octadec-7-ene-4-carbonyl]-amide(94)

Compound 93 (65 mg, 0.12 mmol), DMAP (21 mg, 0.17 mmol) and EDAC (44 mg,0.23 mmol) was dissolved in DMF (0.2 ml). The reaction mixture wasstirred for 6 h at R.T. whereafter cyclopropylsulfonamide (69 mg, 0.57mmol) and DBU (80 μl, 0.57 mmol) was added. After stirring at R.Tovernight the reaction mixture was extracted between EtOAc (80 ml) andaqueous citric add (10%, 2×15 ml). The organic phase was dried overMgSO₄, evaporated on silica and purified twice by chromatography onsilica gel (5% MeOH in dichloromethane→15% MeOH in dichloromethane)which gave a syrup. This syrup was dissolved in a small volumeacetonitrile and precipitated with ethyl ether to give the pure titleproduct (19 mg, 23%). MS (M+H⁺): 673.2

Example 95

1-{[2-Hex-5-enyl-methyl-carbamoyl)-4-(7-methoxy-2-phenyl-quinolin-4-yloxy)-cyclopentanecarbonyl]-amino-2-vinyl-cyclopropanecarboxylicadd ethyl ester (95)

The tert.butyl ester of compound 35 was removed according to theprocedure described in Example 39. The afforded acid (850 mg, 1.56mmol), N-methyl hex-6-enylamine hydrochloride (380 mg, 2.5 mmol) anddiisopropylethylamine (2.3 ml, 13.4 mmol) was dissolved in DMF (60 mL)and cooled to 0° C. After 30 min. HATU (0.76 mg, 2.0 mmol) was added andthe flask was removed from the ice-bath. The reaction was followed withTLC. After 2 h the reaction mixture was added to 5% citric acid andextracted three times with ethyl acetate. The organic phase was driedover sodium sulphate and evaporated under reduced pressure. The crudeproduct was purified by silica gel chromatography which gave the titleproduct (820 mg, 82%.

Example 96

17-(7-Methoxy-2-phenyl-quinolin-4-yloxy)-3-methyl-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carboxylicacid ethyl ester (96)

Compound 95 (648 mg, 1.01 mmol) was dissolved in degassed dichloroethane(500 mL). Hoveyda-Grubbs catalyst 2:nd generation (35 mg, 0.055 mmol)was added and the mixture was refluxed under argon atmosphere overnight.The reaction mixture was evaporated on silica and purified bychromatography on silica gel (30% EtOAc in toluene→50% EtOAc in toluene)to give the pure title product (230 mg mg, 37%). MS (M+H⁺): 612.8

Example 97

17-(7-Methoxy-2-phenyl-quinolin-4-yloxy)-13-methyl-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carboxylicadd ethyl ester (97)

Compound 96 (260 mg, 0.42 mmol) was dissolved in 1,4-dioxan (20 mL), 1.0M Lithium hydroxide (6.0 ml) was added and the mixture was stirred atroom temperature overnight and then for six hours at 60° C. The mixturewas added to 5% citric acid and extracted 3 times with ethyl acetate.The organic phase was dried over sodium sulphate and evaporated underreduced pressure. The crude product was purified by silica gelchromatography with DCM and 5% MeOH which gave the title product (130mg, 53%). MS (M+H): 584.7

Example 98

Cyclopropanesulphonic acid[17-(7-methoxy-2-phenyl-quinolin-4-yloxy)-13-methyl-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*octadec-7ene-4-carbonyl]-amide(98)

Compound 97 (58.3 mg, 0.1 mmol), DMAP (18.3 mg, 0.15 mmol) and EDAC(38.7 mg, 0.2 mmol) was dissolved in DMF (1.0 mL). The reaction mixturewas stirred overnight at R.T. whereafter cyclopropylsulphonamide (60.5mg, 0.5 mmol) and DBU (76 μg, 0.5 mmol) was added. After stirring at R.Tovernight the reaction mixture was added to 5% citric acid and extractedthree times with ethyl acetate. The organic phase was dried over sodiumsulphate and evaporated. The afforded residue was purified two times bysilica gel chromatography which gave the title product (20 mg). MS (M+H)687.8.

Example 99

[4-Cyclopropanesulphonylaminocarbonyl-17(7-methoxy-phenyl-quinolin-4-yloxy)-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6]octadec-7-en-13-yl]-carbamicacid tert.butyl ester (99)

N′-Hex-5-en-(E)-ylidene-hydrazinecarboxylic acid tert.butyl ester wasprepared according to the procedure described in Example 48 and 47 butstarting from hex-5-en-ol instead of hept-6-en-ol. Compound 35 wastreated as described in Example 48 but using the above describedN′-Hex-5-en-(E)-ylidene-hydrazinecarboxylic acid tert.butyl esterinstead of the corresponding hept-6-en derivative followed bymacrocyclisation as described in Example 49 and hydrolysis of the ethylester as described in Example 50 gave the acid. The afforded acid (58mg, 0.0848 mmol) was dissolved in dry DMF (7 mL) and DIEA was added dropwise during one minute. The solution was stirred at room temperature for1 h prior to the addition of a solution containingcyclopropylsulfonamide (41 mg, 0.338 mmol), DMAP (41.3 mg, 0.338 mmol)and DBU (50 μL, 0.338 mmol) in dry DMF (1.5 mL). The solution wasstirred at room temperature for 5 days. The solution was diluted withEtOAc (50 mL) and washed with sat NaHCO₃. The aqueous phase wasextracted with DCM. The combined organic layers were dried, concentratedand subjected to purification by HPLC, which gave the title compound asa white solid (14.3 mg, 0.018 mmol), Purity by HPLC>95%, M+H⁺ 788.3.

Example 100

Cyclopropanesulphonicacid[13-amino-17-(7-methoxy-2-phenyl-quinolin-4-yloxy)-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carbonyl]-amidetrifluoroacetic add salt (100)

Compound 99 (2.4 mg, 0.00304 mmol) was kept in TFA-DCM 1:2 (3 mL) atroom temperature for 60 min. Toluene (3 mL) was added. The sample wasco-evaporated to dryness to afford the title compound (2.1 mg, 0.0028mmol) Purity by HPLC>95%. M+H⁺ 688.3.

Example 101

3-Oxo-2-oxa-bicyclo[2.2.1]heptane-5-carboxylic acidhex-5-enyl-methylamide (101)

To HATU (2.17 g, 5.7 mmol) and N-methyl hex-5-enylamine hydrochloride(8.47 mmol) in 5 mL DMF, under argon in an ice bath, were added1R,4,5R-3-oxo-2-oxa-bicyclo[2.2.1]heptane-5-carboxylic acid (835.6 mg,5.35 mmol) in 11 mL DMF followed by DIEA (2.80 mL, 16 mmol). Afterstirring for 40 min, the mixture was stirred at rt for 5 h. The solventwas evaporated, the residue dissolved in EtOAc (70 mL) and washed withsaturated NaHCO₃ (10 mL). The aqueous phase was extracted with EtOAc(2×25 mL). The organic phases were combined, washed with saturated NaCl(20 mL), dried over Na₂SO₄, and evaporated. Flash column chromatography(150 g silica gel, 2/1 EtOAc—petroleum ether (PE), TLC detection byaqueous KMnO4, Rf 0.55 in 4/1 EtOAc—PE) gave the compound as a yellowoil (1.01 g, 75%).

Example 102

4-Hydroxycyclopentane-1,2-dicarboxylic acid1-[(1-cyclopropanesulphonylaminocarbonyl-2-vinylcyclopropyl)-amide]2-(hex-5-enyl-methylamide (102)

LiOH solution (0.15M, 53 mL, 8 mmol) was added to the lactone amide 101(996 mg, 3.96 mmol) in an ice bath and stirred for 1 h. The mixture wasacidified to pH 2-3 with 1N HC and evaporated, co-evaporated withtoluene several times, and dried under vacuum overnight(1R,2S)-cyclopropanesulfonic acid(1-amino-2-vinyl-cyclopropanecarbonyl)amide hydrochloride (421 mmol) andHATU (1.78 g, 4.68 mmol) were added. The mixture was cooled in an icebath under argon, DMF (25 mL) and then DIEA (2.0 mL, 11.5 mmol) wereadded. After stirring for 30 min, the mixture was stirred at rt for 3 h.After evaporation of solvent, the residue was dissolved in EtOAc (120mL), washed successively with 0.5 N HCl (20 mL) and saturated NaCl (2×20mL), and dried over Na₂SO₄. Flash column chromatography (200 g YMCsilica gel, 2-4% MeOH in CH₂Cl₂ gave white solids (1.25 g, 66%).

Example 103

Cyclopropanesulphonic acid(17-hydroxy-13-methyl-2,14-dioxo-3,13-diazatricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carbonyl)-amide(103)

The cyclopentanol 102 (52.0 mg, 0.108 mmol) was dissolved in 19 mL1,2-dichloroethane (bubbled with argon prior to use). The Hoveyda-Grubbs2^(nd) generation catalyst (6.62 mg, 10 mole %) was dissolved in DCE(2×0.5 mL) and added. The green solution was bubbled with Ar for 1 min.Aliquots (4 mL each) were transferred Into five 2 to 5-mL microwavetubes. To the last tube was added 0.8 mL rinsing with solvent. Each tubewas heated by microwave (rt to 160° C. In 5 min). All aliquots werecombined and the solvent evaporated. Flash column chromatography (silicagel, 3-7% MeOH in CH₂Cl₂) gave 24.39 mg solids (Rf 0.28 in 10%MeOH—CH₂C₁ with two spots). The solids were combined with a 9.86-mgsample and subjected to a second chromatography (2-8% MeOH in EtOAc) togive cream solids (23 mg) with 80% of the desired compound (26% yield).

Example 104

Cyclopropanesulphonic add{17-[2-(4-isopropylthiazol-2-yl)-7-methoxyquinolin-4-yloxy]-13-methyl-2,14-dioxo-3,13-diazatricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carbonyl)-amide(104)

DIAD (22 uL, 0.11 mmol) was added to a mixture of the metathesis product103 (23 mg), 2-(4-isopropyl-1,3-thiazol-2-yl)-7-methoxyquinolin-4-ol (24mg, 0.08 mmol), and PPh₃(30 mg, 0.11 mmol) in 1 mL dry THF, in an icebath. The mixture was stirred at rt overnight and then evaporated. Theresidue (1.2 mL of a 1.5-mL MeCN solution) was purified by prep-HPLC(Hypercarb 7 uL 100×212 mm, 40% to 99% aqueous MeCN in 10 min) to give3.18 mg MV062308 as cream solids (13% yield). 1H NMR (DMSO-d6) δ ppm:major rotamer 0.99 (m, 2H), 1.11 (m, 2H), 1.20-1.30 (m, 2H), 1.37 and1.38 (2d, J=7.0 Hz, 6H), 1.46-1.58 (m, 2H), 1.70 (m, 1H), 1.85 (m, 1H),1.90 (dd, J=8.5, 6.0 Hz, 1H), 2.08 (br, 1H), 2.26 (m, 1H), 2.38 (m, 1H),2.52-2.82 (m, 3H), 2.90-2.97 (m, 2H), 3.06 (s, 3H), 3.21 (m, 1H),3.40-3.56 (m, 2H) 3.97 (s, 3H), 4.60 (m, 1H), 5.04 (m, 1H), 5.41 (br,1H), 5.66 (m, 1H), 7.16 (m), 7.58 (br), 8.02 (m), 10.92 (s, 1H)

Example 105

N-{4-[4-Cyclopropanesulphonylaminocarbonyl-13-methyl-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6]octadec-7-en-17-yloxy)-7methoxy-quinolin-2-yl]-thiazol-2-yl}-3,3dimethylbutyramide(106)

Treatment of compound 103 with4-hydroxy-7-methoxy-2-[2-(2,2-dimethylbutanoyl)aminothiazol-4-yl]quinolineas described hi Example 104 gave the title compound.

LCMS: retention time 2.30 in gradient 30%-80% B in 3 min(flow. 0.8mL/min, UV 220 nm, ACE C8 3×5 mm; mobile phase A 10 mM NH₄Ac in 90% H₂O,B 10 mM NH₄Ac in 90% ACN), (M+1)⁺=807.

Example 106

1-{[2-(Hex-5-enyl-methyl-carbamoyl)-4-hydroxy-cyclopentanecarbonyl]-amino}-2-vinyl-cyclopropanecarboxylicacid ethyl ester (106)

Reaction of compound 101 as described hi example 102 but using1-amino-2-vinylcyclopropanecarboxylic acid ethyl ester instead of(1R,2S)-cyclopropanesulfonic acid(1-amino-2-vinyl-cyclopropanecarbonyl)amide hydrochloride gave the titlecompound.

Example 107

1-{(4-(4-Bromo-benzensulphonyloxy-2-(hex-5-enyl-methyl-carbamoyl)-cyclopentanecarbonyl-amino}2-vinyl-cyclopropanecarboxylicadd ethyl ester (107)

Compound 106 (115 mg, 0.286 mmol) was dissolved in toluene 5 ml anddichloromethane 1 ml. DABCO (2.2.2-diazobicyclooctane) (96 mg, 0.857mmol, 3 eq.) was added to the solution, followed by addition of BsCl(109 mg, 0.428 mmol, 1.5 eq). The reaction was stirred at roomtemperature overnight, diluted with toluene (+10% ethyl acetate), washedwith saturated sodium bicarbonate, brine, dried over sodium sulphate andevaporated. The desired product was obtained by column chromatography(eluent EtOAc) R_(f) 0.25). Conversion 80%. Yield 106 mg.

Example 108

17-(4-Bromo-benzensulphonyloxy)-13-methyl-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carboxylicacid ethyl ester (108)

Compound 107 (106 mg, 0.169 mmol) was dissolved in dichloromethane (40ml) and degassed by bubbling nitrogen through the solution for 20 min.Hoveyda-Grubbs catalyst 1st generation (10 mg, 0.017 mmol, 10 mol %) wasthen added and the mixture was refluxed under nitrogen atmosphereovernight. The reaction mixture was then cooled down to room temperatureand MP-TMT palladium scavenger (approx 100 mg) was added and stirred for2.5 h. The scavenger was removed by filtration and washed with 50 ml ofdichloromethane. The solution obtained was concentrated by rotaryevaporation. The crude was purified by column chromatography (EtOAc) togive 61 mg of product. Yield 60%.

Example 109

17-[2-(2-isopropylamino-triazol-4-yl)-7-methoxy-quinolin-4-yloxy]-13-methyl-2,14-dioxo-3,13-diazatricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carboxylicacid ethyl ester (109)

2-(Isopropylamino-thiazol-4-yl)-7-methoxy-quinolin-4-ol (220 mg, 0.7mmol) (prepared as described in WO 00/59929) was dissolved in 7 mil ofNMP (N-methyl pyrrolidinone), one spoon of Cs₂CO₃ was added, stirred at60° C. for 1.5 h. Then compound 108 (150 mg, 0.24 mmol) was added. Thereaction mixture was stirred at 80° C. overnight. Was diluted withchloroform and washed with sodium bicarbonate, brine. Water phases wereback-extracted with chloroform. The combined organic layers were driedover sodium sulphate and evaporated. The crude product was purified bypreparative HPLC (Gilson) (MeOH—H₂O, 65%) to give 21 mg of product(yield 13%) as well as 12 mg of isomer.

Example 110

17-[2-(2-Isopropylamino-thiazol-4-yl)-7-methoxy-quinolin-4-yloxy]-13-methyl-2,14-dioxo-3,13-diaza-tricyclo[13.3.0.0*4,6*]octadec-7-ene-4-carboxylicacid (110)

To the solution of the ester 109 (21 mg, 0.031 mmol) in a mixture of THF(02 ml) and methanol (0.3 ml) was added solution of LiOH (4 mg, 0.17mmol) in 0.15 ml water. The resulting mixture was stirred at 60° C. for3.5 h. After cooling to room temperature, acetic acid was added (30 eq).The mixture was co-evaporated with toluene. The residue was distributedbetween chloroform and water, the water phase was extracted withchloroform 3 times, the organic phases were combined, dried over sodiumsulphate and evaporated which gave 20 mg of pure product (yield 99%).

Example 111

Cyclopropanesulphonic acid{17-[2-(2-isopropylamino-thiazol-4-yl)-7-methoxy-quinolin-4-yloxy]-13-methyl-2,14-dioxo-3,13-diaza-tricyclo[(13.3.0.0*4,6*]octadec-7-ene-4-carbonyl}amide (111)

The acid 110 (20 mg, 0.15 mmol), DMAP (28 mg, 0.225 mmol) and EDAC (58mg, 0.3 mmol) was dissolved in DMF (1.5 mL). The reaction mixture wasstirred overnight at R.T. whereafter cyclopropylsulphonamide (91 mg,1.125 mmol) and DBU (114 μL, 0.75 mmol) was added. After stirring at RTovernight the reaction mixture was added to 5% citric add and extractedthree times with chloroform. The organic phase was dried over sodiumsulphate and evaporated. The afforded residue was purified bypreparative HPLC to give the title product (5.6 mg) (yield 24%).

Assays

The compounds of the invention are conveniently assayed for activityagainst the NS3 protease of flavivirus such as HCV using conventional invitro (enzyme) assays or cell culture assays.

A useful assay is the Bartenshlager replicon assay disclosed in EP1043399. An alternative replicon assay is described in WO 03064416.

A convenient enzyme assay involving the inhibition of full-lengthhepatitis C NS3 is essentially as described in Poliakov, 2002 ProtExpression & Purification 25 363 371.

Briefly, the hydrolysis of a depsipeptide substrate,Ac-DED(Edans)EEAbu_(ψ)[COO]ASK(Dabcyl)-NH₂ (AnaSpec, San José, USA), ismeasured spectrofluorometrically in the presence of a peptide cofactor,KKGSVVIVGRIVLSGK, as described by Landro, 1997 Biochem 36 9340-9348. Theenzyme (1 nM) is incubated in a buffer such as 50 mM HEPES, pH 7.5, 10mM DTT, 40% glycerol, 0.1% n-octyl-β-D-glucoside, with 25 μM cofactorand inhibitor at say 30° C. for 10 min, whereupon the reaction isinitiated by addition of substrate, typically 0.5 μM substrate.Inhibitors are typically dissolved in DMSO, sonicated for 30 s andvortexed. The solutions are generally stored at −20° betweenmeasurements.

An alternative enzyme assay is described in WO 0399316 and employs anHCV NS3/4A protease complex FRET peptide assay. The purpose of this invitro assay is to measure the inhibition of HCV NS3 protease complexes,derived from the BMS, H77C or J4168 strains, as described below, bycompounds of the present invention. This assay provides an indication ofhow effective compounds of the present invention would be in inhibitingHCV proteolytic activity.

Serum is taken from an HCV-infected patient. An engineered full-lengthcDNA template of the HCV genome (BMS strain) was constructed from DNAfragments obtained by reverse transcription-PCR (RT-PCR) of serum RNAand using primers selected on the basis of homology between othergenotype I a strains. From the determination of the entire genomesequence, a genotype I a was assigned to the HCV isolate according tothe classification of Simmonds et al. (See P Simmonds, K A Rose, SGraham, SW Chan, F McOmish, B C Dow, E A Follett, P L Yap and H Marsden,J. Clin. Microbiol., 31(6), 1493-1503 (1993)). The amino add sequence ofthe nonstructural region, NS2-58, was shown to be >97% identical to HCVgenotype Ia (H77C) and 87% identical to genotype Ib (J4L6S). Theinfectious clones, H77C (I a genotype) and J4L6S (b genotype) can beobtained from R. Purcell (NIH) and the sequences are published inGenbank (AAB6703, see Yanagi, M., Purcell, R. H., Emerson, S. U. andBukh. Proc. Natl. Acad. Sd. U.S.A. 94 (16) 8738-8743 (1997); AF054247,see Yanagi, M., St Claire, M., Shapiro, M., Emerson, S. U., Purcell, R.H. and Bukhj, Virology 244 (1), 161 (1998)).

The BMS, H77C and J4L6S strains are conventional for production ofrecombinant NS3/4A protease complexes. DNA encoding the recombinant HCVNS3/4A protease complex (amino acids 1027 to 1711) for these strainswere manipulated as described by P. Gallinari at al. (see Gallinari P,Paolini C, Brennan D, Nardi C, Steinkuhler C, De Francesco R.Biochemistry. 38(17):562032, (1999)). Briefly, a three-lysinesolubilizing tall was added at the 3′-end of the 3 0 NS4A coding region.The cysteine in the P1 position of the NS4A-NS4B cleavage site (aminoacid 1711) was changed to a glycine to avoid the proteolytic cleavage ofthe lysine tag. Furthermore, a cysteine to serine mutation can beintroduced by PCR at amino acid position 1454 to prevent the autolyticcleavage in the NS3 helices domain. The variant DNA fragment can becloned in the pET21b bacterial expression vector (Novagen) and the NS34Acomplex can be expressed in Escherichia coli strain BL21 (DE3)(invitrogen) following the protocol described by P. Gallinari et a. (seeGallinari P, Brennan D, Nardi C, Brunetti M, Tomei L, Steinkuhler C, DeFrancesco R., J Virol. 72(8):6758-69 (1998)) with modifications.Briefly, NS3/4A expression can be induced with 0.5 mM isopropyl beta-Dthiogalactopyranoside (IPTG) for 22 hr at 20′C. A typical fermentation(I0 I) yields approximately 80 g of wet cell paste. The cells areresuspended in lysis buffer (IO mL/g) consisting of 25 mMN-(2Hydroxyethyl)Piperazine-N′-(2-Ethane Sulfonic acid) (HEPES), pH7.5,20% glycerol, 500 mM Sodium Chloride (NaCl), 0.5% Triton-X100, I ug/mLlysozyme, 5 mM Magnesium Chloride (MgCl₂), I ug/mL Dnasel, 5 mMbeta-Mercaptoethanol (BME), Protease inhibitor—EthylenediamineTetraacetic acid (EDTA) free (Roche), homogenized and incubated for 20mine at VC. The homogenate is sonicated and clarified byultra-centrifugation at 235000 g for 1 hr at 4° C.

Imidazole is added to the supernatant to a final concentration of 15 mMand the pH adjusted to 8. The crude protein extract is loaded on aNickel Nitrilotriacetic add (Ni-NTA) column pre-equilibrated with bufferB (25n-tM 2.0 HEPES, pH8 20% glycerol, 500 mM NaCl, 0.5% Triton-XIOO, 15mM imidazole, 5 mM BME). The sample is loaded at a flow rate of 1mL/min. The column is washed with 15 column volumes of buffer C (same asbuffer B except with 02% Triton-X100). The protein s eluted with 5column volumes of buffer D (same as buffer C except with 200 mMimidazole).

NS3/4A protease complex-containing fractions are pooled and loaded on adesalting column Superdex-S200 pre-equilibrated with buffer D (25 MMHEPES, pH7.5, 20% glycerol, 300 mM NaCl, 0.2% Triton-XIOO, IO mM BME).Sample is loaded at a flow rate of ImUmin. NS3/4A protease complex3 0containing fractions are pooled and concentrated to approximately 0.5mg/mL The purity of the NS3/4A protease complexes, derived from the BMS,H77C and J4L6S strains, are typically judged to be greater than 90% bySDS-PAGE and mass spectrometry analyses.

The enzyme is generally stored at −80′C, thawed on ice and diluted priorto use in assay buffer. The substrate used for the NS3/4A proteaseassay, is conveniently RET S 1 (Resonance Energy Transfer DepsipeptideSubstrate; AnaSpec, inc. cat #22991)(FRET peptide), described by Talianiat al. in Anal. Biochem. 240(2):6067 (1996). The sequence of thispeptide is loosely based on the NS4A/NS4B natural cleavage site exceptthere is an ester linkage rather than an amide bond at the cleavagesite. The peptide substrate is incubated with one of the threerecombinant NS3/4A complexes, in the absence or presence of a compoundof the present invention, and the formation of fluorescent reactionproduct was followed in real time using a Cytofluor Series 4000. Usefulreagents are as follow: HEPES and Glycerol (Ultrapure) can be obtainedfrom GIBCO-BRL. Dimethyl Sulfoxide (DMSO) is obtained from Sigma.Beta-Mercaptoethanol is obtained from Bio Rad.

Assay buffer: 50 m.M HEPES, pH7.5; 0.15M NaCl; 0.1% Triton; 15%Glycerol; 10 mM BME. Substrate: 2 uM final concentration (from a 2 mMstock 2 0 solution in DMSO stored at −20′C). HCV NS3/4A type Ia (Ib),2-3 nM final concentration (from a 5 uM stock solution in 25 mM HEPES,pH7.5, 20% glycerol, 300 m.M NaCl, 0.2% Triton-X100, 10 mM BME). Forcompounds with potencies approaching the assay limit, the assay can bemade more sensitive by adding 50 ug/mL BSA to the assay buffer and/orreducing the end protease concentration to 300 pM.

The assay is conveniently performed in a 96-well polystyrene black platefrom Falcon. Each well contains 25 ul NS3/4A protease complex in assaybuffer, 50 ul of a compound of the present invention in 10% DMSO/assaybuffer and 25 ul substrate in assay buffer. A control (no compound) isalso prepared on the same assay plate. The enzyme complex is mixed withcompound or control solution, typically for 1 min before initiating theenzymatic reaction by the addition of substrate. The assay plate isgenerally read immediately using a spectrophotometer such as a CytofluorSeries 4000 (Perspective Biosystems). The instrument is conveniently setto read an emission of 340 nm and excitation of 490 nm at 25′C.Reactions are generally followed for approximately 15 minutes.

The percent inhibition can be calculated with the following equation.100.−[(dF _(inh) /dF _(con))×100]where dF is the change in fluorescence over the linear range of thecurve. A nonlinear curve fit is applied to the inhibition-concentrationdata, and the 50% effective concentration (IC₅₀) is calculated by theuse software such as Excel XI-fit software using the equation:y=A+((B−A)(1+((C/x)^D))).

Enzyme assays conveniently utilize a fluorescence resonance energytransfer (FRET) principle to generate a spectroscopic response to an HCVNS3 serine protease catalyzed NS4A/48 cleavage event. The activity istypically measured in a continuous fluorometric assay using anexcitation wavelength of 355 nm and emission wavelength of 500 nm. Theinitial velocity may be determined from 10 minutes continuous reading ofincreased fluorescence intensities as a result of the NS3 proteasecatalyzed cleavage event.

An alternative enzyme assay can be carried out as follows:

Materials

Recombinant HCV NS3 full length enzyme can be prepared as shown inPoliakov at at Protein Expression & purification 25 (2002) 363-371. TheNS4A cofactor conveniently has an amino acid sequence ofKKGSVVIVGRIVLSGK (commercially available), generally prepared as a 10 mMstock solution in DMSO. The FRET-substrate(Ac-Asp-Glu-Asp(EDANS)-Glu-Glu-Abu-ψ-(COO)Ala-Ser-Lys(DABCYL)-NH2,MW1548.60 can be purchased from AnaSpec RET S1, CA. USA) and istypically prepared as a 1.61 mM stock solution in DMSO. Aliquots (50μl/tube) should be wrapped with aluminum foil to protect from directlight and stored in −20° C.

Reference compound-1, N-1725 with a sequence ofAcAsp-D-Gla-Leu-Ile-Cha-Cys, MW 830.95 may be purchased from BACHEM,Switzerland and is generally prepare as a 2 mM stock solution in DMSOand stored in aliquots in −20° C.

1M HEPES buffer may be purchased from invitrogen Corporation, storage at20° C. Glycerol may be purchased from Sigma, 99% purity.

CHAPS, 3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate: may bepurchased from Research Organics, Cleveland, Ohio 44125, USA. MW614.90DTT, DL-Dithiothreitol (Cleland Reagent DL-DT) 99% purity, MW.1542Storage: +4° C.

DMSO may be purchased from SDS, 13124 Peypin, France. 99.5% purity.

TRIS, ultra pure (TRIS-(hydroxymethylaminomethane), may be purchasedfrom ICN Biomedicals Inc.

Sodium Chloride, may be obtained from KEBOlab AB.

N-dodecyl-β-maltoside, minimum 98%, may be purchased from Sigma, storage−20° C.

Equipment

Microtiter plates (white clinlplate, ThermoLab Systems cat no. 9502890

Eppendorf pipettes

Biohit pipette, multi dosing

Ascent fluorimeter, flterpai ex 355 nm, em500 nm

Method

Experimental Procedure:

10 mM stack solutions of the compounds are made in DMSO. The stocksolutions are stored in room temperature while testing and placed in−20° C. at long-time storage.

Assay Buffer A:

50 mM HEPES buffer, pH=7.5, 40% Glycerol, 0.1% CHAPS

Storage: room temperature.

10 mM DTT (stored in aliquots at −20° C. and added fresh at eachexperiment)

Assay Buffer B:

25 mM TRIS pH7.5, 0.15 M NaCl, 10% glycerol, 0.05%n-dodecyl-β-D-maltoside

5 mM DTT (stored in aliquots at −20° C. and added fresh at eachexperiment)

Assay Sequence:

Preparation of Reaction Buffer (for One Date. 100Reactions)(Buffer A)

1. Prepare 9500 μl assay buffer (HEPES, pH=7.5, 40% glycerol and 0.1%CHAPS in de ionized water. Add DTT giving a final concentration of 10 mM(freshly prepared for every run).

2. Thaw rapidly the NS3 protease

3. Add 13.6 μl NS3 protease and 13.8 μl NS4A peptide and mix properly.Leave the mixture for 15 minutes in room temperature.

4. Place the enzyme stock solution back into liquid nitrogen or −80° C.as soon as possible.

Preparation of Reaction Buffer (for One Plate, 100Reactions) (Buffer B)

5. Prepare 9500 μl assay buffer (TRIS, pH=7.5, 0.16 M NaCl, 0.5 mM EDTA,10% glycerol and 0.05% n-dodecyl β-D-maltoside in de ionized water. AddDTT giving a final concentration of 5 mM (freshly prepared for everyrun).

6. Thaw the NS3 protease rapidly.

7. Add 27.2 μl NS3 protease and 13.6 μl NS4A peptide and mix property.Leave the mixture for 15 minutes in room temperature.

8. Place the enzyme stock solution back into liquid nitrogen or −80° C.as soon as possible.

Preparation of Inhibitor/Reference Compound

Make a dilution series of the inhibitors in DMSO to 100× the finalconcentrations 10, 1, 0.1, 0.01 and 0.001 μM. The final DM80concentration in 100 μl total reaction volume is 1%.

Make a dilution series of the reference compound, N-1725 in DMSO to 100×the final concentrations 120, 60, 30, 15, 7.5 and 3.75 nM.

Eight enzyme control wells are needed for every run.

Blank wells contain 95 μL buffer (without NS3 PR), 1 μL DMSO and 5 μLsubstrate.

Preparation of FRET Substrate

Dilute the substrate stock solution (1.61 mM) with assay buffer to 40 μMworking solution. Avoid exposure to light.

Assay Sequence

Use 96-well cliniplate, the total assay volume per well is 100 μl.

1. Add 95 μL of assay buffer to each well

2. Add 1 μl inhibitor/reference compound

3. Pre incubate for 30 minutes at room temperature

4. Start the reaction by adding 5 μL 40 μM substrate solution (finalconcentration 2 μM)

5. Read continuously for 20 minutes at ex=355 nm and em=500 nm,monitoring the increased fluorescence per minute.

6. Plot the progression curve (within linear range, 8-10 time points)and determine the slope as an initial velocity with respect to eachindividual inhibitor concentration.

7. Calculate % inhibition with respect to enzyme control.

Treatment of Results

The result is expressed as % inhibition at a certain concentration(screen) or as a Ki value in nM or μM.

Calculation of % inhibition: The initial velocity is determined from 10minutes continuous reading of increased fluorescence intensities as aresult of the NS3 protease catalyzed cleavage event. The change in slopefor the inhibitor compared to the enzyme control gives the % inhibitionat a certain concentration.

Calculation of Ki: All inhibitors are treated as if they follow therules of competitive inhibition. The IC₅₀ value is calculated from theinhibition values of a series of inhibitor concentrations. Thecalculated value it used in the following equation: K_(i)=IC₅₀/(1+S/Km)

Plotting of the graph is done by help of two calculation programs:Grafit and Graphpad

Various compounds of the invention exemplified above displayed IC₅₀s inthe range 1 nM to 6.9 micromolar and ED₅₀s in the sub-micromolar tomicromolar range in the above assays.

Drug Escape Resistance Pattern and Rate

Replicon cultures in microtitre plates can be used to determineresistance development rates and to select out drug escape mutants. Thecompounds being tested are added at concentrations around their ED₅₀using, say, 8 duplicates per concentration. After the appropriatereplicon incubation period the protease activity in the supernatant orlysed cells is measured.

The following procedure is followed at subsequent passages of thecultures. Virus produced at the concentration of test compoundshowing >50% of the protease activity of untreated infected cells (SIC,Starting Inhibitory Concentration) are passaged to fresh repliconcultures. An aliquot, say, 15 μl supernatant from each of the eightduplicates are transferred to replicon cells without the test compound(control) and to cells with test compound at the same concentration, andadditionally two respectively fivefold higher concentrations. (See thetable below) When the viral component of replicon propagation (forexample as measured by HCV protease activity) is permitted at thehighest non-toxic concentration (5-40 μM), 2-4 parallel wells arecollected and expanded to give material for sequence analysis andcross-wise resistance.

Key:

Viral Growth Emitted

Virus production inhibited

Pass 1 Pass 2 Pass 3 Pass 4 Pass 5 SIC No compound SIC No compound  5 ×SIC SIC 25 × SIC  5 × SIC No compound 25 × SIC  5 × SIC No compound  25× SIC  5 × SIC 125 × SIC  25 × SIC 125 × SIC

Alternative methods for assessing activity on drug escape mutantsinclude the preparation of mutant enzyme bearing the distinctivemutation for use in standard Ki determinations as shown above. Forexample WO 04/039970 describes constructions allowing access to HCVproteases bearing the 155, 156 and/or 168 drug escape mutants arisingfrom the selective pressure of BILN-2061 and VX-950. Such constructs canthen be engineered into replicon vectors in place of the wild typeprotease, thereby allowing ready assessment in a cellular assay, ofwhether a given compound is active against a give drug escape mutant.

P450 Metabolism

The metabolism of compounds of the invention through the main isoformsof the human cytochrome system P450 are conveniently determined inbaculovirus infected insect cells transfected with human cytochrome P450cDNA (supersomes) Gentest Corp. Woburn USA.

The test compounds at concentrations 0.5, 5 and 50 μM are incubated induplicate in the presence of supersomes overexpressing variouscytochrome P450 isoforms, including CYP1A2+P450 reductase, CYP2A6+P450reductase, CYP2C9-Arg 144+P450 reductase, CYP2C19+P450 reductase,CYP2D6-Val 374+P450 reductase and CYP3A4+P 450 reductase. Incubatescontain a fixed concentration of cytochrome P450 (eg 50 pmoles) and areconducted over 1 hour. The involvement of a given isoform in themetabolism of the test compound is determined by UV HPLCchromatographically measuring the disappearance of parent compound.

The invention claimed is:
 1. A compound of formula VIga:

wherein A is C(═O)OR¹ or C(═O)NHSO₂R²; wherein R₁ is hydrogen orC₁-C₆alkyl; and R₂ is C₁-C₆alkyl, C₀-C₆alkylcarbocyclyl, orC₀-C₃alkylheterocyclyl, each of which is optionally substituted with 1to 3 substituents which are each independently halo, oxo, nitrile,azido, nitro, C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl,NH₂C(═O)—, Y—NRaRb, Y—O—Rb, YC(═O)Rb, Y—(C═O)NRaRb, Y—NRaC(═O)Rb,Y—NHSOpRb, Y—S(═O)pRb, YS(═O)pNRaRb, Y—C(═O)ORb, or Y—NRaC(═O)ORb; Y isindependently a bond or C₁-C₃alkylene; p is independently 1 or 2; Ra isindependently H or C₁-C₃alkyl; Rb is independently H, C₁-C₆alkyl,C₀-C₃alkylcarbocyclyl, or C₀-C₃alkylheterocyclyl; Rd is H or C₁-C₃alkyl;q′ is 0 and k is 1; Rz is H, or together with the asterisked carbonforms an olefinic bond; Rq is H or C₁-C₆alkyl; W is —O— or —S—; R₈ is aring system containing 1 or 2 saturated, partially saturated orunsaturated rings, each of which has 4-7 ring atoms and each of whichhas 0 to 4 hetero atoms selected from S, O, and N, the ring system beingoptionally spaced from W by a C₁-C₃alkyl group; any of which R⁸ groupscan be optionally mono, di, or tri substituted with R⁹, wherein R₉ isindependently halo, oxo, nitrile, azido, nitro, C₁-C₆alkyl,C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl, NH₂C(═O)—, YNRaRb,Y—O—Rb, Y—C(═O)Rb, Y—(C═O)NRaRb, Y—NRaC(═O)Rb, Y-NHSOpRb, YS(═O)pRb,Y—S(═O)pNRaRb, Y—C(═O)ORb, or Y—NRaC(═O)ORb; wherein said carbocyclyl orheterocyclyl moiety is optionally substituted with R¹⁰; wherein R₁₀ isC₁-C₆alkyl, C₃-C₇cycloalkyl, C₁-C₆alkoxy, amino, sulfonyl, (C₁-C₃alkyl)sulfonyl, NO₂, OH, SH, halo, haloalkyl, carboxyl, or amido; J is asingle 3 to 10-membered saturated or partially unsaturated alkylenechain, which chain is optionally interrupted by one to three heteroatomsthat are each independently —O—, —S—, or —NR¹²—, and wherein 0 to 3carbon atoms in the chain are optionally substituted with R¹⁴; R₁₂ is H,C₁-C₆ alkyl, C₃-C₆cycloalkyl, or COR¹³; R₁₃ is C₁-C₆alkyl,C₀-C₃alkylcarbocyclyl, or C₀-C₃alkylheterocyclyl; each R¹⁴ isindependently H, C₁-C₆alkyl, C₁-C₆haloalkyl, C₁-C₆alkoxy, hydroxyl,halo, amino, oxo, thio, or C₁-C₆ thioalkyl; G is —O—, —NRy-, or NRjNRj-;Ry is H or C₁-C₃alkyl; Rj is H; U is ═O or is absent; R₁₆ is H; or R¹⁶is C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, or C₀-C₃alkylheterocyclyl, any ofwhich can be substituted with halo, oxo, nitrile, azido, nitro,C₁-C₆alkyl, C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl, NH₂CO—,Y—NRaRb, Y—O—Rb, YC(═O)Rb, Y—(C═O)NRaRb, Y—NRaC(═O)Rb, Y-NHSOpRb,Y—S(═O)pRb, YS(═O)pNRaRb, Y—C(═O)ORb, or Y—NRaC(═O)ORb; or apharmaceutically acceptable salt thereof.
 2. The compound according toclaim 1, wherein R² is C₁-C₆alkyl, C₀-C₃alkylaryl, C₃-C₇cycloalkyl, orphenyl, each of which is optionally substituted with 1 to 3 substituentswhich are each halo, oxo, nitrile, azido, nitro, C₁-C₆alkyl,C₀-C₃alkylcarbocyclyl, C₀-C₃alkylheterocyclyl, NH₂C(═O)—, Y—NRaRb,Y—O—Rb, YC(═O)Rb, Y—(C═O)NRaRb, Y—NRaC(═O)Rb, Y-NHSOpRb, Y—S(═O)pRb,YS(═O)pNRaRb, Y—C(═O)Orb, or Y—NRaC(═O)ORb.
 3. The compound according toclaim 1, wherein said compound has the partial structure that is:


4. The compound according to claim 3, wherein Rq is C₁-C₃ alkyl.
 5. Thecompound according to claim 1, wherein A is C(═O)OR¹, wherein R¹ is H orC₁-C₆ alkyl.
 6. The compound according to claim 1, wherein W is —O—. 7.The compound according to claim 1 wherein R⁸ is optionally substitutedC₀-C₃alkylcarbocyclyl or optionally substituted C₀-C₃alkylheterocyclyl.8. The compound according to claim 7, wherein R⁸ is 1-naphthylmethyl,2-naphthylmethyl, benzyl, 1-naphthyl, 2-napthyl, or quinolinyl any ofwhich is unsubstituted, mono, or disubstituted with R⁹ as defined. 9.The compound according to claim 8, wherein R⁸ is:

wherein R^(9a) is C₁-C₆alkyl; C₁-C₆alkoxy; thioC₁-C₃alkyl; aminooptionally substituted with C₁-C₆alkyl; C₀-C₃alkylaryl;C₀-C₃alkylheteroaryl, or C₀-C₃alkylheterocyclyl, said aryl, heteroarylor heterocycle being optionally substituted with R¹⁰ wherein R¹⁰ isC₁-C₆alkyl; C₀-C₃alkyl; C₃-C₇cycloalkyl; C₁-C₆alkoxy; amino optionallymono- or di-substituted with C₁-C₆alkyl; amide; or C₁-C₃alkylamide; andR^(9b) is C₁-C₆alkyl, C₁-C₆-alkoxy, amino, di(C₁-C₃alkyl)amino,(C₁-C₃alkyl)amide, NO₂, OH, halo, trifluoromethyl, or carboxyl.
 10. Thecompound according to claim 9, wherein R^(9a) is aryl or heteroaryl,either of which is optionally substituted with R¹⁰ as defined.
 11. Thecompound according to claim 9, wherein R^(9a) is:

wherein R¹⁰ is H; C₁-C₆alkyl; C₀-C₃alkylcycloalkyl; amino optionallymono- or di-substituted with C₁-C₆alkyl; amido; or (C₁-C₃alkyl)amide.12. The compound according to claim 9, wherein R^(9a) is optionallysubstituted phenyl.
 13. The compound according to claim 9, whereinR^(9b) is C₁-C₆-alkoxy.
 14. The compound according to claim 1, wherein Jis a 4 to 7-membered saturated or unsaturated, all carbon alkylenechain.
 15. The compound according to claim 1, wherein R¹⁶ is H,C₁-C₆alkyl, or C₃-C₆cycloalkyl.
 16. A pharmaceutical compositioncomprising a compound as defined in claim 1 and a pharmaceuticallyacceptable carrier.
 17. The pharmaceutical composition of claim 16,further comprising an additional HCV antiviral.
 18. The pharmaceuticalcomposition of claim 17, wherein the additional HCV antiviral is anucleoside analogue polymerase inhibitor, a protease inhibitor,ribavirin, or interferon.
 19. A method of treating hepatitis C virus(HCV) infection comprising administering to an individual afflicted withor at risk of HCV infection, an effective amount of a compound ofclaim
 1. 20. The method of claim 19, further comprising administering tothe individual, an additional HCV antiviral.
 21. The method of claim 20,wherein the additional HCV antiviral is a nucleoside analogue polymeraseinhibitor, a protease inhibitor, ribavirin, or interferon.